Protein moonlighting
Encyclopedia
Protein moonlighting is a phenomenon by which a protein
can perform more than one function. Ancestral moonlighting proteins originally possessed a single function but through evolution
, acquired additional functions. Many proteins that moonlight are enzyme
s; others are receptor
s, ion channel
s or chaperones. The most common primary function of moonlighting proteins is enzymatic catalysis
, but these enzymes have acquired secondary non-enyzmatic roles. Some examples of functions of moonlighting proteins secondary to catalysis include signal transduction
, transcriptional regulation
, apoptosis
, motility
, and structural.
Protein moonlighting may occur widely in nature. Protein moonlighting through gene sharing differs from the use of a single gene to generate different protein by alternative RNA splicing
, DNA rearrangement, or post-translational processing
. It is also different from multifunctionality of the protein, in which the protein has multiple domains, each serving a different function. Protein moonlighting by gene sharing means that a gene may acquire and maintain a second function without gene duplication and without loss of the primary function. Such genes are under two or more entirely different selective constraints.
Various techniques have been used to reveal moonlighting functions in proteins. The detection of a protein in unexpected locations within cells, cell types, or tissues may suggest that a protein has a moonlighting function. Furthermore, sequence or structure homology of a protein may be used to infer both primary function as well as secondary moonlighting functions of a protein.
The most well-studied examples of gene sharing are crystallin
s. These proteins, when expressed at low levels in many tissues function as enzymes, but when expressed at high levels in eye tissue, become densely packed and thus form lenses. While the recognition of gene sharing is relatively recent—the term was coined in 1988, after crystallins in chickens and ducks were found to be identical to separately identified enzymes—recent studies have found many examples throughout the living world. Joram Piatigorsky has suggested that many or all proteins exhibit gene sharing to some extent, and that gene sharing is a key aspect of molecular evolution
. The genes encoding crystallins must maintain sequences for catalytic function and transparency maintenance function.
Inappropriate moonlighting is a contributing factor in some genetic diseases, and moonlighting provides a possible mechanism by which bacteria may become resistant to antibiotics.
enzymes. Piatigorsk determined that lens crystallins' conservation and variance is due to other moonlighting functions outside of the lens. Originally Piatigorsky called these proteins "gene sharing" proteins, but the colloquial description moonlighting was subsequently applied to proteins by Constance Jeffery in 1999 to draw a similarity between multitasking proteins and people who work two jobs. The phrase "gene sharing" is ambiguous since it is also used to describe horizontal gene transfer
, hence the phrase "protein moonlighting" has become the preferred description for proteins with more than one function.
through which uni-functional proteins gained the ability to perform multiple functions. With alterations, much of the protein's unused space can provide new functions. Many moonlighting proteins are the result of gene fusion
of two single function genes. Alternatively a single gene can acquire a second function since the active site of the encoded protein typically is small compared to the overall size of the protein leaving considerable room to accommodate a second functional site. In yet a third alternative, the same active site can acquire a second function through mutations of the active site.
The development of moonlighting proteins may be evolutionary favorable to the organism since a single protein can do the job of multiple proteins conserving amino acids and energy required to synthesize these proteins. However there is no universally agreed upon theory that explains why proteins with multiple roles evolved. While using one protein to perform multiple roles seems advantageous because it keeps the genome small, we can conclude that this is probably not the reason for moonlighting because of the large of amount of noncoding DNA
.
. Other proteins fulfill structural, transport, or signaling roles. Furthermore, numerous proteins have the ability to aggregate into supramolecular assemblies
. For example, a ribosome
is made up of 90 proteins and RNA
.
A number of the currently known moonlighting proteins are evolutionarily derived from highly conserved
enzymes, also called ancient enzymes. These enzymes are frequently speculated to have evolved moonlighting functions. Since highly conserved proteins are present in many different organisms, this increases the chance that they would develop secondary moonlighting functions. A high fraction of enzymes involved in glycolysis
, an ancient universal metabolic pathway, exhibit moonlighting behavior. Furthermore it has been suggested that as many as 7 out of 10 proteins in glycolysis and 7 out of 8 enzymes of the tricarboxylic acid cycle exhibit moonlighting behavior.
An example of a moonlighting enzyme is pyruvate carboxylase
. This enzyme catalyzes the carboxylation of pyruvate
into oxaloacetate
, thereby replenishing the tricarboxylic acid cycle
. Surprisingly, in yeast species such as H. polymorpha
and P. pastoris
, pyruvate carboylase is also essential for proper targeting and assembly of the peroxisomal protein alcohol oxidase
(AO). AO, the first enzyme of methanol metabolism, is a homo-octameric flavoenzyme. In wild type cells, this enzyme is present as enzymatically active AO octamers in the peroxisomal
matrix. However, in cells lacking pruvate carboxylase, AO monomers accumulate in the cytosol, indicating that pyruvate carboxylase has a second fully unrelated function in assembly and import. The function in AO import/assembly is fully independent of the enzyme activity of pyruvate carboxylase, because amino acid substitutions can be introduced that fully inactive the enzyme activity of pryuvate carboxylase, without affecting its function in AO assembly and import. Conversely, mutations are known that block the function of this enzyme in import and assembly of AO, but have no effect on the enzymatic activity of the protein.
The E. coli
anti-oxidant thioredoxin
protein is another example of a moonlighting protein. Upon infection with the bacteriophage T7
, E. coli thioredoxin forms a complex with T7 DNA polymerase
, which results in enhanced T7 DNA replication, a crucial step for successful T7 infection. Thioredoxin binds to a loop in T7 DNA polymerase to bind more strongly to the DNA. The anti-oxidant function of thioredoxin is fully autonomous and fully independent of T7 DNA replication, in which the protein most likely fulfills the functional role.
) will function as a protease
by the directed degradation of proteins and in lower temperatures as a chaperone by assisting the non-covalent folding or unfolding and the assembly or disassembly of other macromolecular structures. Furthermore, moonlighting proteins may exhibit different behaviors not only as a result of its location within a cell, but also the type of cell that the protein is expressed in.
Other methods through which proteins may moonlight are by changing their oligomeric
state, altering concentrations of the protein's ligand or substrate, use of alternative binding sites, or finally through phosphorylation
. An example of a protein that displays different function in different oligomeric states is pyruvate kinase
which exhibits metabolic activity as a tetramer and thyroid hormone
–binding activity as a monomer. Changes in the concentrations of ligands or substrates may cause a switch in protein a protein's function. For example, in the presence of low iron concentrations, aconitase
functions as an enzyme while at high iron concentration, aconitase functions as an iron-responsive element-binding protein (IREBP). Proteins may also perform separate functions through the use of alternative binding sites that perform different tasks. An example of this is ceruloplasmin
, a protein that functions as an oxidase in copper metabolism and moonlights as a copper-independent glutathione peroxidase
. Lastly, phosphorylation may sometimes cause a switch in the function of a moonlighting protein. For example, phosphorylation of phosphoglucose isomerase
(PGI) at Ser-185 by protein kinase CK2
causes it to stop functioning as an enzyme, while retaining its function as an autocrine
motility
factor. Hence when a mutation takes place that inactivates a function of a moonlighting proteins, the other function(s) are not necessarily affected.
The crystal structures of several moonlighting proteins, such as I-AniI homing endonuclease
/ maturase and the PutA proline dehydrogenase
/ transcription factor
, have been determined. An analysis of these crystal structures has demonstrated that moonlighting proteins can either perform both functions at the same time, or through conformational change
s, alternate between two states, each of which is able to perform a separate function. For example, the protein DegP plays a role in proteolysis with higher temperatures and is involved in refolding functions at lower temperatures. Lastly, these crystal structures have shown that the second function may negatively affect the first function in some moonlighting proteins. As seen in ƞ-crystallin, the second function of a protein can alter the structure, decreasing the flexibility, which in turn can impair enzymatic activity somewhat.
Various methods have been employed to determine a protein's function including secondary moonlighting functions. For example, the tissue, cellular, or subcellular distribution of a protein may provide hints as to the function. Real-time PCR
is used to quantify mRNA
and hence infer the presence or absence of a particular protein which is encoded by the mRNA within different cell types. Alternatively immunohistochemistry
or mass spectrometry
can be used to directly detect the presence of proteins and determine in which subcellular locations, cell types, and tissues a particular protein is expressed.
Mass spectrometry may be used to detect proteins based on their mass-to-charge ratio
. Because of alternative splicing
and posttranslational modification
, identification of proteins based on the mass of the parent ion alone is very difficult. However tandem mass spectrometry
in which each of the parent peaks is in turn fragmented can be used to unambiguously identify proteins. Hence tandem mass spectrometry is one of the tools used in proteomics
to identify the presence of proteins in different cell types or subcellular locations. While the presence of a moonlighting protein in an unexpected location may complicate routine analyses, at the same time, the detection of a protein in unexpected multiprotein complexes or locations suggests that protein may have a moonlighting function. Furthermore, mass spectrometry may be used to determine if a protein has high expression levels that do not correlate to the enzyme's measured metabolic activity. These expression levels may signify that the protein is performing a different function than previously known.
The structure
of a protein can also help determine its functions. Protein structure in turn may be elucidated with various techniques including X-ray crystallography or NMR
. Dual polarization interferometry may be used to measure changes in protein structure which may also give hints to the protein's function. Finally, application of systems biology
approaches such as interactomics
give clues to a proteins function based on what it interacts with.
In the case of crystallins, the genes must maintain sequences for catalytic function and transparency maintenance function. The abundant lens crystallins have been generally viewed as static proteins serving a strictly structural role in transparency and cataract
. However, recent studies have shown that the lens crystallins are much more diverse than previously recognized and that many are related or identical to metabolic enzymes and stress proteins found in numerous tissues. Unlike other proteins performing highly specialized tasks, such as globin
or rhodopsin
, the crystallins are very diverse and show numerous species differences. Essentially all vertebrate lenses contain representatives of the α and β/γ crystallins, the "Ubiquitous crystallins", which are themselves heterogeneous, and only few species or selected taxonomic groups use entirely different proteins as lens crystallins.This paradox of crystallins being highly conserved in sequence while extremely diverse in number and distribution shows that many crystallins have vital functions outside the lens and cornea, and this multi-functionality of the crystallins is achieved by gene sharing.
. The fact that similar transcriptional factors such as Pax-6, and retinoic acid receptors, regulate different crystalline genes, suggests that lens-specific expression have played a crucial role for recruiting multifunctional protein as crystallins. Crystallin recruitment has occurred both with and without gene duplication, and tandem gene duplication has taken place among some of the crystallins with one of the duplicates specializing for lens expression. Ubiquitous α –crystallins and bird δ –crystallins are two examples.
, cornea
is a transparent, avascular tissue derived from the ectoderm
that is responsible for focusing light onto the retina
. However, unlike lens, cornea depends on the air-cell interface and its curvature for refraction. Early immunology studies have shown that BCP 54 comprises 20–40% of the total soluble protein in bovine cornea. Subsequent studies have indicated that BCP 54 is ALDH3, a tumor and xenobiotic-inducible cytosolic enzyme, found in human, rat, and other mammals.
. The α-crystallins may also play a functional role in the stability and remodeling of the cytoskeleton during fiber cell differentiation in the lens. In cornea, ALDH3 is also suggested to be responsible for absorbing UV-B light.
, it necessarily involves separate functions at the molecular level. A gene could exhibit pleiotropy when single enzyme function affects multiple phenotypic traits
; mutations of a shared gene could potentially affect only a single trait. Gene duplication
followed by differential mutation is another phenomenon thought to be a key element in the evolution of protein function, but in gene sharing, there is no divergence of gene sequence when proteins take on new functions; the single polypeptide takes on new roles while retaining old ones. Alternative splicing
can result in the production of multiple polypeptides (with multiple functions) from a single gene, but by definition, gene sharing involves multiple functions of a single polypeptide.
from genotype
, hampering the study of inherited metabolic disorder
s.
The complex phenotypes of several disorders are suspected to be caused by the involvement of moonlighting proteins. The protein GAPDH
has at least 11 documented functions, one of which includes apoptosis. Excessive apoptosis is involved in many neurodegenerative diseases, such as Huntington's
, Alzheimer's
, and Parkinson's
as well as in brain ischemia
. In one case, GAPDH was found in the degenerated neurons of individuals who had Alzheimer's disease.
Although there is insufficient evidence for definite conclusions, there are well documented examples of moonlighting proteins that play a role in disease. One such disease is tuberculosis
. One moonlighting protein in the bacterium M. tuberculosis
has a function which counteracts the effects of antibiotics. Specifically, M. tuberculosis gains antibiotic resistance
against ciprofloxacin
from overexpression of Glutamate racemase
in vivo
.
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
can perform more than one function. Ancestral moonlighting proteins originally possessed a single function but through evolution
Evolution
Evolution is any change across successive generations in the heritable characteristics of biological populations. Evolutionary processes give rise to diversity at every level of biological organisation, including species, individual organisms and molecules such as DNA and proteins.Life on Earth...
, acquired additional functions. Many proteins that moonlight are enzyme
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
s; others are receptor
Receptor (biochemistry)
In biochemistry, a receptor is a molecule found on the surface of a cell, which receives specific chemical signals from neighbouring cells or the wider environment within an organism...
s, ion channel
Ion channel
Ion channels are pore-forming proteins that help establish and control the small voltage gradient across the plasma membrane of cells by allowing the flow of ions down their electrochemical gradient. They are present in the membranes that surround all biological cells...
s or chaperones. The most common primary function of moonlighting proteins is enzymatic catalysis
Enzyme catalysis
Enzyme catalysis is the catalysis of chemical reactions by specialized proteins known as enzymes. Catalysis of biochemical reactions in the cell is vital due to the very low reaction rates of the uncatalysed reactions....
, but these enzymes have acquired secondary non-enyzmatic roles. Some examples of functions of moonlighting proteins secondary to catalysis include signal transduction
Signal transduction
Signal transduction occurs when an extracellular signaling molecule activates a cell surface receptor. In turn, this receptor alters intracellular molecules creating a response...
, transcriptional regulation
Transcriptional regulation
Transcriptional regulation is the change in gene expression levels by altering transcription rates. -Regulation of transcription:Regulation of transcription controls when transcription occurs and how much RNA is created...
, apoptosis
Apoptosis
Apoptosis is the process of programmed cell death that may occur in multicellular organisms. Biochemical events lead to characteristic cell changes and death. These changes include blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, and chromosomal DNA fragmentation...
, motility
Motility
Motility is a biological term which refers to the ability to move spontaneously and actively, consuming energy in the process. Most animals are motile but the term applies to single-celled and simple multicellular organisms, as well as to some mechanisms of fluid flow in multicellular organs, in...
, and structural.
Protein moonlighting may occur widely in nature. Protein moonlighting through gene sharing differs from the use of a single gene to generate different protein by alternative RNA splicing
Alternative splicing
Alternative splicing is a process by which the exons of the RNA produced by transcription of a gene are reconnected in multiple ways during RNA splicing...
, DNA rearrangement, or post-translational processing
Posttranslational modification
Posttranslational modification is the chemical modification of a protein after its translation. It is one of the later steps in protein biosynthesis, and thus gene expression, for many proteins....
. It is also different from multifunctionality of the protein, in which the protein has multiple domains, each serving a different function. Protein moonlighting by gene sharing means that a gene may acquire and maintain a second function without gene duplication and without loss of the primary function. Such genes are under two or more entirely different selective constraints.
Various techniques have been used to reveal moonlighting functions in proteins. The detection of a protein in unexpected locations within cells, cell types, or tissues may suggest that a protein has a moonlighting function. Furthermore, sequence or structure homology of a protein may be used to infer both primary function as well as secondary moonlighting functions of a protein.
The most well-studied examples of gene sharing are crystallin
Crystallin
In anatomy, a crystallin is a water-soluble structural protein found in the lens and the cornea of the eye accounting for the transparency of the structure. It has also been identified in other places such as the heart, and in aggressive breast cancer tumors....
s. These proteins, when expressed at low levels in many tissues function as enzymes, but when expressed at high levels in eye tissue, become densely packed and thus form lenses. While the recognition of gene sharing is relatively recent—the term was coined in 1988, after crystallins in chickens and ducks were found to be identical to separately identified enzymes—recent studies have found many examples throughout the living world. Joram Piatigorsky has suggested that many or all proteins exhibit gene sharing to some extent, and that gene sharing is a key aspect of molecular evolution
Molecular evolution
Molecular evolution is in part a process of evolution at the scale of DNA, RNA, and proteins. Molecular evolution emerged as a scientific field in the 1960s as researchers from molecular biology, evolutionary biology and population genetics sought to understand recent discoveries on the structure...
. The genes encoding crystallins must maintain sequences for catalytic function and transparency maintenance function.
Inappropriate moonlighting is a contributing factor in some genetic diseases, and moonlighting provides a possible mechanism by which bacteria may become resistant to antibiotics.
Discovery
The first observation of a moonlighting protein was made in the late 1980s by Joram Piatigorsky and Graeme Wistow during their research of crystallinCrystallin
In anatomy, a crystallin is a water-soluble structural protein found in the lens and the cornea of the eye accounting for the transparency of the structure. It has also been identified in other places such as the heart, and in aggressive breast cancer tumors....
enzymes. Piatigorsk determined that lens crystallins' conservation and variance is due to other moonlighting functions outside of the lens. Originally Piatigorsky called these proteins "gene sharing" proteins, but the colloquial description moonlighting was subsequently applied to proteins by Constance Jeffery in 1999 to draw a similarity between multitasking proteins and people who work two jobs. The phrase "gene sharing" is ambiguous since it is also used to describe horizontal gene transfer
Horizontal gene transfer
Horizontal gene transfer , also lateral gene transfer , is any process in which an organism incorporates genetic material from another organism without being the offspring of that organism...
, hence the phrase "protein moonlighting" has become the preferred description for proteins with more than one function.
Evolution
It is believed that moonlighting proteins came about by means of evolutionEvolution
Evolution is any change across successive generations in the heritable characteristics of biological populations. Evolutionary processes give rise to diversity at every level of biological organisation, including species, individual organisms and molecules such as DNA and proteins.Life on Earth...
through which uni-functional proteins gained the ability to perform multiple functions. With alterations, much of the protein's unused space can provide new functions. Many moonlighting proteins are the result of gene fusion
Fusion gene
A fusion gene is a hybrid gene formed from two previously separate genes. It can occur as the result of a translocation, interstitial deletion, or chromosomal inversion...
of two single function genes. Alternatively a single gene can acquire a second function since the active site of the encoded protein typically is small compared to the overall size of the protein leaving considerable room to accommodate a second functional site. In yet a third alternative, the same active site can acquire a second function through mutations of the active site.
The development of moonlighting proteins may be evolutionary favorable to the organism since a single protein can do the job of multiple proteins conserving amino acids and energy required to synthesize these proteins. However there is no universally agreed upon theory that explains why proteins with multiple roles evolved. While using one protein to perform multiple roles seems advantageous because it keeps the genome small, we can conclude that this is probably not the reason for moonlighting because of the large of amount of noncoding DNA
Noncoding DNA
In genetics, noncoding DNA describes components of an organism's DNA sequences that do not encode for protein sequences. In many eukaryotes, a large percentage of an organism's total genome size is noncoding DNA, although the amount of noncoding DNA, and the proportion of coding versus noncoding...
.
Functions
Many proteins catalyze a chemical reactionChemical reaction
A chemical reaction is a process that leads to the transformation of one set of chemical substances to another. Chemical reactions can be either spontaneous, requiring no input of energy, or non-spontaneous, typically following the input of some type of energy, such as heat, light or electricity...
. Other proteins fulfill structural, transport, or signaling roles. Furthermore, numerous proteins have the ability to aggregate into supramolecular assemblies
Supramolecular assembly
A supramolecular assembly or "supermolecule" is a well defined complex of molecules held together by noncovalent bonds. While a supramolecular assembly can be simply composed of two molecules , it is more often used to denote larger complexes of molecules that form sphere-, rod-, or sheet-like...
. For example, a ribosome
Ribosome
A ribosome is a component of cells that assembles the twenty specific amino acid molecules to form the particular protein molecule determined by the nucleotide sequence of an RNA molecule....
is made up of 90 proteins and RNA
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....
.
A number of the currently known moonlighting proteins are evolutionarily derived from highly conserved
Conserved sequence
In biology, conserved sequences are similar or identical sequences that occur within nucleic acid sequences , protein sequences, protein structures or polymeric carbohydrates across species or within different molecules produced by the same organism...
enzymes, also called ancient enzymes. These enzymes are frequently speculated to have evolved moonlighting functions. Since highly conserved proteins are present in many different organisms, this increases the chance that they would develop secondary moonlighting functions. A high fraction of enzymes involved in glycolysis
Glycolysis
Glycolysis is the metabolic pathway that converts glucose C6H12O6, into pyruvate, CH3COCOO− + H+...
, an ancient universal metabolic pathway, exhibit moonlighting behavior. Furthermore it has been suggested that as many as 7 out of 10 proteins in glycolysis and 7 out of 8 enzymes of the tricarboxylic acid cycle exhibit moonlighting behavior.
An example of a moonlighting enzyme is pyruvate carboxylase
Pyruvate carboxylase
Pyruvate carboxylase is an enzyme of the ligase class that catalyzes the irreversible carboxylation of pyruvate to form oxaloacetate .It is an important anaplerotic reaction that creates oxaloacetate from pyruvate...
. This enzyme catalyzes the carboxylation of pyruvate
Pyruvic acid
Pyruvic acid is an organic acid, a ketone, as well as the simplest of the alpha-keto acids. The carboxylate ion of pyruvic acid, CH3COCOO−, is known as pyruvate, and is a key intersection in several metabolic pathways....
into oxaloacetate
Oxaloacetic acid
Oxaloacetic acid is an organic compound with the chemical formula C4H4O5 or HOOC---COOH. It also has other names ....
, thereby replenishing the tricarboxylic acid cycle
Citric acid cycle
The citric acid cycle — also known as the tricarboxylic acid cycle , the Krebs cycle, or the Szent-Györgyi-Krebs cycle — is a series of chemical reactions which is used by all aerobic living organisms to generate energy through the oxidization of acetate derived from carbohydrates, fats and...
. Surprisingly, in yeast species such as H. polymorpha
Hansenula polymorpha
Ogataea polymorpha is a methylotrophic yeast with unusual characteristics. It is used as a protein factory for pharmaceuticals.Ogataea polymorpha belongs to a limited number of methylotrophic yeast species – yeasts that can grow on methanol...
and P. pastoris
Pichia pastoris
Pichia pastoris is a species of methylotrophic yeast. Pichia is widely used for protein expression using recombinant DNA techniques. Hence it is used in biochemical and genetic research in academia and the biotechnical industry....
, pyruvate carboylase is also essential for proper targeting and assembly of the peroxisomal protein alcohol oxidase
Alcohol oxidase
In enzymology, an alcohol oxidase is an enzyme that catalyzes the chemical reactionThus, the two substrates of this enzyme are primary alcohol and O2, whereas its two products are aldehyde and H2O2....
(AO). AO, the first enzyme of methanol metabolism, is a homo-octameric flavoenzyme. In wild type cells, this enzyme is present as enzymatically active AO octamers in the peroxisomal
Peroxisome
Peroxisomes are organelles found in virtually all eukaryotic cells. They are involved in the catabolism of very long chain fatty acids, branched chain fatty acids, D-amino acids, polyamines, and biosynthesis of plasmalogens, etherphospholipids critical for the normal function of mammalian brains...
matrix. However, in cells lacking pruvate carboxylase, AO monomers accumulate in the cytosol, indicating that pyruvate carboxylase has a second fully unrelated function in assembly and import. The function in AO import/assembly is fully independent of the enzyme activity of pyruvate carboxylase, because amino acid substitutions can be introduced that fully inactive the enzyme activity of pryuvate carboxylase, without affecting its function in AO assembly and import. Conversely, mutations are known that block the function of this enzyme in import and assembly of AO, but have no effect on the enzymatic activity of the protein.
The E. coli
Escherichia coli
Escherichia coli is a Gram-negative, rod-shaped bacterium that is commonly found in the lower intestine of warm-blooded organisms . Most E. coli strains are harmless, but some serotypes can cause serious food poisoning in humans, and are occasionally responsible for product recalls...
anti-oxidant thioredoxin
Thioredoxin
Thioredoxin is a class of small redox proteins known to be present in all organisms. It plays a role in many important biological processes. In humans, it is encoded by the TXN gene. Loss-of-function mutation of either of the two human thioredoxin genes is lethal at the four-cell stage of the...
protein is another example of a moonlighting protein. Upon infection with the bacteriophage T7
T7 phage
Bacteriophage T7 is a bacteriophage capable of infecting susceptible bacterial cells. It infects most strains of Escherichia coli Bacteriophage T7 is a bacteriophage capable of infecting susceptible bacterial cells. It infects most strains of Escherichia coli Bacteriophage T7 is a bacteriophage...
, E. coli thioredoxin forms a complex with T7 DNA polymerase
T7 DNA polymerase
The T7 DNA polymerase of the T7 bacteriophage is a DNA-dependent DNA polymerase responsible for the fast rate of T7 phage DNA replication in vivo. The polymerase consists of a 1:1 complex of the viral T7 gene 5 protein and the E...
, which results in enhanced T7 DNA replication, a crucial step for successful T7 infection. Thioredoxin binds to a loop in T7 DNA polymerase to bind more strongly to the DNA. The anti-oxidant function of thioredoxin is fully autonomous and fully independent of T7 DNA replication, in which the protein most likely fulfills the functional role.
| Kingdom Kingdom (biology) In biology, kingdom is a taxonomic rank, which is either the highest rank or in the more recent three-domain system, the rank below domain. Kingdoms are divided into smaller groups called phyla or divisions in botany... | | Protein | | Organism | | Function | |
|---|---|---|---|---|
| primary | moonlighting | |||
| Animal Animal Animals are a major group of multicellular, eukaryotic organisms of the kingdom Animalia or Metazoa. Their body plan eventually becomes fixed as they develop, although some undergo a process of metamorphosis later on in their life. Most animals are motile, meaning they can move spontaneously and... |
||||
| Aconitase Aconitase Aconitase is an enzyme that catalyses the stereo-specific isomerization of citrate to isocitrate via cis-aconitate in the tricarboxylic acid cycle, a non-redox-active process.- Function :... |
H. sapiens | TCA cycle enzyme | Iron homeostasis | |
| ATF2 | H. sapiens | Transcription factor | DNA damage respons | |
| Crystallins | Various | Lens structural protein | Various enzyme | |
| Cytochrome c Cytochrome c The Cytochrome complex, or cyt c is a small heme protein found loosely associated with the inner membrane of the mitochondrion. It belongs to the cytochrome c family of proteins. Cytochrome c is a highly soluble protein, unlike other cytochromes, with a solubility of about 100 g/L and is an... |
Various | Energy metabolism | Apoptosis | |
| DLD Dihydrolipoamide dehydrogenase Dihydrolipoamide dehydrogenase , also known as dihydrolipoyl dehydrogenase, mitochondrial, is an enzyme that in humans is encoded by the DLD gene. DLD is a flavoprotein enzyme that degrades lipoamide, and produces dihydrolipoamide.... |
H. sapiens | Energy metabolism | Protease | |
| ERK2 | H. sapiens | MAP kinase | Transcriptional repressor | |
| ESCRT ESCRT ESCRT refers to a series of cytosolic protein complexes called ESCRT-0, ESCRT-I, ESCRT-II, and ESCRT-III.-Endocytosis and Receptor Down-Regulation:... -II complex |
D. melanogaster Drosophila melanogaster Drosophila melanogaster is a species of Diptera, or the order of flies, in the family Drosophilidae. The species is known generally as the common fruit fly or vinegar fly. Starting from Charles W... |
Endosomal protein sorting | Biocoid mRNA localization | |
| STAT3 STAT3 Signal transducer and activator of transcription 3 also known as STAT3 is a transcription factor which in humans is encoded by the STAT3 gene.- Function :The protein encoded by this gene is a member of the STAT protein family... |
M. musculus | Transcription factor | Electron transport chain | |
| Plant Plant Plants are living organisms belonging to the kingdom Plantae. Precise definitions of the kingdom vary, but as the term is used here, plants include familiar organisms such as trees, flowers, herbs, bushes, grasses, vines, ferns, mosses, and green algae. The group is also called green plants or... |
||||
| Hexokinase Hexokinase A hexokinase is an enzyme that phosphorylates a six-carbon sugar, a hexose, to a hexose phosphate. In most tissues and organisms, glucose is the most important substrate of hexokinases, and glucose-6-phosphate the most important product.... |
A. thaliana Arabidopsis thaliana Arabidopsis thaliana is a small flowering plant native to Europe, Asia, and northwestern Africa. A spring annual with a relatively short life cycle, arabidopsis is popular as a model organism in plant biology and genetics... |
Glucose metabolism | Glucose signaling | |
| Presenilin Presenilin Presenilins are a family of related multi-pass transmembrane proteins that function as a part of the gamma-secretase intramembrane protease complex... |
P. patens Physcomitrella patens Physcomitrella patens is a moss used as a model organism for studies on plant evolution, development and physiology.-Model organism:... |
γ-secretase | Cystoskeletal function | |
| Fungus Fungus A fungus is a member of a large group of eukaryotic organisms that includes microorganisms such as yeasts and molds , as well as the more familiar mushrooms. These organisms are classified as a kingdom, Fungi, which is separate from plants, animals, and bacteria... |
||||
| Aconitase Aconitase Aconitase is an enzyme that catalyses the stereo-specific isomerization of citrate to isocitrate via cis-aconitate in the tricarboxylic acid cycle, a non-redox-active process.- Function :... |
S. cerevisiae Saccharomyces cerevisiae Saccharomyces cerevisiae is a species of yeast. It is perhaps the most useful yeast, having been instrumental to baking and brewing since ancient times. It is believed that it was originally isolated from the skin of grapes... |
TCA cycle enzyme | mtDNA stability | |
| Aldolase Aldolase Aldolase A is an enzyme that catalyses a reverse aldol reaction: The substrate, fructose 1,6-bisphosphate is broken down into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate . This reaction is a part of glycolysis. Three aldolase isozymes , encoded by three different genes, are... |
S. cerevisiae | Glycolytic enzyme | V-ATPase assembly | |
| Arg5,6 | S. cerevisiae | Arginine biosynthesis | Transcriptional control | |
| Enolase Enolase Enolase, also known as phosphopyruvate dehydratase, is a metalloenzyme responsible for the catalysis of the conversion of 2-phosphoglycerate to phosphoenolpyruvate , the ninth and penultimate step of glycolysis. Enolase belongs to the class Lyase. Enolase can also catalyze the reverse reaction,... |
S. cerevisiae | Glycolytic enzyme |
|
|
| Galactokinase Galactokinase Galactokinase is an enzyme that facilitates the phosphorylation of α-D-galactose to galactose 1-phosphate at the expense of one molecule of ATP. Galactokinase catalyzes the second step of the Leloir pathway, a metabolic pathway found in most organisms for the catabolism of β-D-galactose to glucose... |
K. lactis Kluyveromyces lactis Kluyveromyces lactis is a Kluyveromyces yeast commonly used for genetic studies and industrial applications. Its name comes from the ability to assimilate lactose and convert it into lactic acid.- Use :... |
Galactose catabolism enyzme | Induction galactose genes | |
| Hal3 | S. cerevisiae | Halotolerance determinant | Coenzyme A biosynthesis | |
| HSP60 HSP60 Heat shock proteins are generally responsible for preventing damage to proteins in response to high levels of heat. Heat shock proteins are classified into six major families based on their molecular mass: small HSPs, HSP40, HSP60, HSP70, HSP90, and HSP110... |
S. cerevisiae | Mitochondrial chaperone | Stabilization active DNA ori's | |
| Phosphofructokinase Phosphofructokinase Phosphofructokinase-1 is the most important regulatory enzyme of glycolysis. It is an allosteric enzyme made of 4 subunits and controlled by many activators and inhibitors... |
P. pastoris Pichia pastoris Pichia pastoris is a species of methylotrophic yeast. Pichia is widely used for protein expression using recombinant DNA techniques. Hence it is used in biochemical and genetic research in academia and the biotechnical industry.... |
Glycolytic enzyme | Autophagy peroxisomes | |
| Pyruvate carboxylase Pyruvate carboxylase Pyruvate carboxylase is an enzyme of the ligase class that catalyzes the irreversible carboxylation of pyruvate to form oxaloacetate .It is an important anaplerotic reaction that creates oxaloacetate from pyruvate... |
H. polymorpha Hansenula polymorpha Ogataea polymorpha is a methylotrophic yeast with unusual characteristics. It is used as a protein factory for pharmaceuticals.Ogataea polymorpha belongs to a limited number of methylotrophic yeast species – yeasts that can grow on methanol... |
Anaplerotic enzyme | Assembly of alcohol oxidase | |
| Vhs3 | S. cerevisiae | Halotolerance determinant | Coenzyme A biosynthesis | |
| Prokaryote Prokaryote The prokaryotes are a group of organisms that lack a cell nucleus , or any other membrane-bound organelles. The organisms that have a cell nucleus are called eukaryotes. Most prokaryotes are unicellular, but a few such as myxobacteria have multicellular stages in their life cycles... s |
||||
| Aconitase Aconitase Aconitase is an enzyme that catalyses the stereo-specific isomerization of citrate to isocitrate via cis-aconitate in the tricarboxylic acid cycle, a non-redox-active process.- Function :... |
M. tuberculosis Mycobacterium tuberculosis Mycobacterium tuberculosis is a pathogenic bacterial species in the genus Mycobacterium and the causative agent of most cases of tuberculosis . First discovered in 1882 by Robert Koch, M... |
TCA cycle enzyme | Iron-responsive protein | |
| CYP170A1 | S. coelicolor Streptomyces coelicolor Streptomyces coelicolor is a soil-dwelling Gram-positive bacterium that belongs to the genus Streptomyces.-Usage in biotechnology:Strains of S. coelicolor produce various antibiotics, including actinorhodin, methylenomycin, undecylprodigiosin, and perimycin. Certain strains of S. coelicolor can... |
Albaflavenone synthase | Terpene synthase | |
| Enolase Enolase Enolase, also known as phosphopyruvate dehydratase, is a metalloenzyme responsible for the catalysis of the conversion of 2-phosphoglycerate to phosphoenolpyruvate , the ninth and penultimate step of glycolysis. Enolase belongs to the class Lyase. Enolase can also catalyze the reverse reaction,... |
S. pneumoniae Streptococcus pneumoniae Streptococcus pneumoniae, or pneumococcus, is Gram-positive, alpha-hemolytic, aerotolerant anaerobic member of the genus Streptococcus. A significant human pathogenic bacterium, S... |
Glycolytic enzyme | Plasminogen binding | |
| GroEL GroEL GroEL belongs to the chaperonin family of molecular chaperones, and is found in a large number of bacteria. It is required for the proper folding of many proteins. To function properly, GroEL requires the lid-like cochaperonin protein complex GroES... |
E. aerogenes Enterobacter aerogenes Enterobacter aerogenes is a Gram-negative, oxidase negative, catalase positive, citrate positive, indole negative, rod-shaped bacterium.... |
Chaperone | Insect toxin | |
| Glutamate racemase Glutamate racemase In enzymology, glutamate racemase is an enzyme that catalyzes the chemical reactionHence, this enzyme RacE has one substrate, L-glutamate, and one product, D-glutamate.... (MurI) |
E. coli Escherichia coli Escherichia coli is a Gram-negative, rod-shaped bacterium that is commonly found in the lower intestine of warm-blooded organisms . Most E. coli strains are harmless, but some serotypes can cause serious food poisoning in humans, and are occasionally responsible for product recalls... |
cell wall biosynthesis | gyrase inhibition | |
| Thioredoxin Thioredoxin Thioredoxin is a class of small redox proteins known to be present in all organisms. It plays a role in many important biological processes. In humans, it is encoded by the TXN gene. Loss-of-function mutation of either of the two human thioredoxin genes is lethal at the four-cell stage of the... |
E. coli | Anti-oxidant | T7 DNA polymerase subunit | |
| Protist Protist Protists are a diverse group of eukaryotic microorganisms. Historically, protists were treated as the kingdom Protista, which includes mostly unicellular organisms that do not fit into the other kingdoms, but this group is contested in modern taxonomy... |
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| Aldolase Aldolase Aldolase A is an enzyme that catalyses a reverse aldol reaction: The substrate, fructose 1,6-bisphosphate is broken down into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate . This reaction is a part of glycolysis. Three aldolase isozymes , encoded by three different genes, are... |
P. vivax Plasmodium vivax Plasmodium vivax is a protozoal parasite and a human pathogen. The most frequent and widely distributed cause of recurring malaria, P. vivax is one of the four species of malarial parasite that commonly infect humans. It is less virulent than Plasmodium falciparum, which is the deadliest of the... |
Glycolytic enzyme | Host-cell invasion | |
Mechanisms
In many cases, the functionality of a protein not only depends on its structure, but also its location. For example, a single protein may have one function when found in the cytoplasm of a cell, a different function when interacting with a membrane, and yet a third function if excreted from the cell. This property of moonlighting proteins is known as "differential localization". For example, in higher temperatures DegP (HtrAHtrA serine peptidase 2
Serine protease HTRA2, mitochondrial is an enzyme that in humans is encoded by the HTRA2 gene.-Interactions:HtrA serine peptidase 2 has been shown to interact with MAPK14, XIAP and BIRC2.-External Links:...
) will function as a protease
Protease
A protease is any enzyme that conducts proteolysis, that is, begins protein catabolism by hydrolysis of the peptide bonds that link amino acids together in the polypeptide chain forming the protein....
by the directed degradation of proteins and in lower temperatures as a chaperone by assisting the non-covalent folding or unfolding and the assembly or disassembly of other macromolecular structures. Furthermore, moonlighting proteins may exhibit different behaviors not only as a result of its location within a cell, but also the type of cell that the protein is expressed in.
Other methods through which proteins may moonlight are by changing their oligomeric
Oligomer
In chemistry, an oligomer is a molecule that consists of a few monomer units , in contrast to a polymer that, at least in principle, consists of an unlimited number of monomers. Dimers, trimers, and tetramers are oligomers. Many oils are oligomeric, such as liquid paraffin...
state, altering concentrations of the protein's ligand or substrate, use of alternative binding sites, or finally through phosphorylation
Phosphorylation
Phosphorylation is the addition of a phosphate group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes....
. An example of a protein that displays different function in different oligomeric states is pyruvate kinase
Pyruvate kinase
Pyruvate kinase is an enzyme involved in glycolysis. It catalyzes the transfer of a phosphate group from phosphoenolpyruvate to ADP, yielding one molecule of pyruvate and one molecule of ATP.-Reaction:The reaction with pyruvate kinase:...
which exhibits metabolic activity as a tetramer and thyroid hormone
Thyroid hormone
The thyroid hormones, thyroxine and triiodothyronine , are tyrosine-based hormones produced by the thyroid gland primarily responsible for regulation of metabolism. An important component in the synthesis of thyroid hormones is iodine. The major form of thyroid hormone in the blood is thyroxine ,...
–binding activity as a monomer. Changes in the concentrations of ligands or substrates may cause a switch in protein a protein's function. For example, in the presence of low iron concentrations, aconitase
Aconitase
Aconitase is an enzyme that catalyses the stereo-specific isomerization of citrate to isocitrate via cis-aconitate in the tricarboxylic acid cycle, a non-redox-active process.- Function :...
functions as an enzyme while at high iron concentration, aconitase functions as an iron-responsive element-binding protein (IREBP). Proteins may also perform separate functions through the use of alternative binding sites that perform different tasks. An example of this is ceruloplasmin
Ceruloplasmin
Ceruloplasmin is a ferroxidase enzyme that in humans is encoded by the CP gene.Ceruloplasmin is the major copper-carrying protein in the blood, and in addition plays a role in iron metabolism. It was first described in 1948...
, a protein that functions as an oxidase in copper metabolism and moonlights as a copper-independent glutathione peroxidase
Glutathione peroxidase
Glutathione peroxidase is the general name of an enzyme family with peroxidase activity whose main biological role is to protect the organism from oxidative damage...
. Lastly, phosphorylation may sometimes cause a switch in the function of a moonlighting protein. For example, phosphorylation of phosphoglucose isomerase
Glucose-6-phosphate isomerase
-Other functions:There is evidence that phosphoglucose isomerase acts as a molecular messenger. It is produced and secreted by white blood cells, and acts to regulate the growth of several different cell types.-Pathology:...
(PGI) at Ser-185 by protein kinase CK2
Casein kinase 2
The Casein kinase 2 is a serine/threonine-selective protein kinase that is a tetramer of two alpha subunits and two beta subunits. The alpha subunits have the catalytic kinase domain...
causes it to stop functioning as an enzyme, while retaining its function as an autocrine
Autocrine signalling
Autocrine signaling is a form of signalling in which a cell secretes a hormone or chemical messenger that binds to autocrine receptors on the same cell, leading to changes in the cell...
motility
Motility
Motility is a biological term which refers to the ability to move spontaneously and actively, consuming energy in the process. Most animals are motile but the term applies to single-celled and simple multicellular organisms, as well as to some mechanisms of fluid flow in multicellular organs, in...
factor. Hence when a mutation takes place that inactivates a function of a moonlighting proteins, the other function(s) are not necessarily affected.
The crystal structures of several moonlighting proteins, such as I-AniI homing endonuclease
Homing endonuclease
The homing endonucleases are a type of restriction enzymes typically encoded by introns or inteins. They act on the cellular DNA of the cells that synthesize them, in the opposite alleles of the genes that encode them.- Origin and mechanism :...
/ maturase and the PutA proline dehydrogenase
Proline dehydrogenase
In enzymology, a proline dehydrogenase is an enzyme that catalyzes the chemical reactionThus, the two substrates of this enzyme are L-proline and acceptor, whereas its two products are -1-pyrroline-5-carboxylate and reduced acceptor....
/ transcription factor
Transcription factor
In molecular biology and genetics, a transcription factor is a protein that binds to specific DNA sequences, thereby controlling the flow of genetic information from DNA to mRNA...
, have been determined. An analysis of these crystal structures has demonstrated that moonlighting proteins can either perform both functions at the same time, or through conformational change
Conformational change
A macromolecule is usually flexible and dynamic. It can change its shape in response to changes in its environment or other factors; each possible shape is called a conformation, and a transition between them is called a conformational change...
s, alternate between two states, each of which is able to perform a separate function. For example, the protein DegP plays a role in proteolysis with higher temperatures and is involved in refolding functions at lower temperatures. Lastly, these crystal structures have shown that the second function may negatively affect the first function in some moonlighting proteins. As seen in ƞ-crystallin, the second function of a protein can alter the structure, decreasing the flexibility, which in turn can impair enzymatic activity somewhat.
Identification methods
Moonlighting proteins have usually been identified by chance because there is no clear procedure to identify secondary moonlighting functions. Despite such difficulties, the number of moonlighting proteins that have been discovered is rapidly increasing. Furthermore, moonlighting proteins appear to be abundant in all kingdoms of life.Various methods have been employed to determine a protein's function including secondary moonlighting functions. For example, the tissue, cellular, or subcellular distribution of a protein may provide hints as to the function. Real-time PCR
Real-time polymerase chain reaction
In molecular biology, real-time polymerase chain reaction, also called quantitative real time polymerase chain reaction or kinetic polymerase chain reaction , is a laboratory technique based on the PCR, which is used to amplify and simultaneously quantify a targeted DNA molecule...
is used to quantify mRNA
Messenger RNA
Messenger RNA is a molecule of RNA encoding a chemical "blueprint" for a protein product. mRNA is transcribed from a DNA template, and carries coding information to the sites of protein synthesis: the ribosomes. Here, the nucleic acid polymer is translated into a polymer of amino acids: a protein...
and hence infer the presence or absence of a particular protein which is encoded by the mRNA within different cell types. Alternatively immunohistochemistry
Immunohistochemistry
Immunohistochemistry or IHC refers to the process of detecting antigens in cells of a tissue section by exploiting the principle of antibodies binding specifically to antigens in biological tissues. IHC takes its name from the roots "immuno," in reference to antibodies used in the procedure, and...
or mass spectrometry
Mass spectrometry
Mass spectrometry is an analytical technique that measures the mass-to-charge ratio of charged particles.It is used for determining masses of particles, for determining the elemental composition of a sample or molecule, and for elucidating the chemical structures of molecules, such as peptides and...
can be used to directly detect the presence of proteins and determine in which subcellular locations, cell types, and tissues a particular protein is expressed.
Mass spectrometry may be used to detect proteins based on their mass-to-charge ratio
Mass-to-charge ratio
The mass-to-charge ratio ratio is a physical quantity that is widely used in the electrodynamics of charged particles, e.g. in electron optics and ion optics. It appears in the scientific fields of lithography, electron microscopy, cathode ray tubes, accelerator physics, nuclear physics, Auger...
. Because of alternative splicing
Alternative splicing
Alternative splicing is a process by which the exons of the RNA produced by transcription of a gene are reconnected in multiple ways during RNA splicing...
and posttranslational modification
Posttranslational modification
Posttranslational modification is the chemical modification of a protein after its translation. It is one of the later steps in protein biosynthesis, and thus gene expression, for many proteins....
, identification of proteins based on the mass of the parent ion alone is very difficult. However tandem mass spectrometry
Tandem mass spectrometry
Tandem mass spectrometry, also known as MS/MS or MS2, involves multiple steps of mass spectrometry selection, with some form of fragmentation occurring in between the stages.-Tandem MS instruments:...
in which each of the parent peaks is in turn fragmented can be used to unambiguously identify proteins. Hence tandem mass spectrometry is one of the tools used in proteomics
Proteomics
Proteomics is the large-scale study of proteins, particularly their structures and functions. Proteins are vital parts of living organisms, as they are the main components of the physiological metabolic pathways of cells. The term "proteomics" was first coined in 1997 to make an analogy with...
to identify the presence of proteins in different cell types or subcellular locations. While the presence of a moonlighting protein in an unexpected location may complicate routine analyses, at the same time, the detection of a protein in unexpected multiprotein complexes or locations suggests that protein may have a moonlighting function. Furthermore, mass spectrometry may be used to determine if a protein has high expression levels that do not correlate to the enzyme's measured metabolic activity. These expression levels may signify that the protein is performing a different function than previously known.
The structure
Protein structure
Proteins are an important class of biological macromolecules present in all organisms. Proteins are polymers of amino acids. Classified by their physical size, proteins are nanoparticles . Each protein polymer – also known as a polypeptide – consists of a sequence formed from 20 possible L-α-amino...
of a protein can also help determine its functions. Protein structure in turn may be elucidated with various techniques including X-ray crystallography or NMR
Protein nuclear magnetic resonance spectroscopy
Nuclear magnetic resonance spectroscopy of proteins is a field of structural biology in which NMR spectroscopy is used to obtain information about the structure and dynamics of proteins. The field was pioneered by Richard R. Ernst and Kurt Wüthrich, among others...
. Dual polarization interferometry may be used to measure changes in protein structure which may also give hints to the protein's function. Finally, application of systems biology
Systems biology
Systems biology is a term used to describe a number of trends in bioscience research, and a movement which draws on those trends. Proponents describe systems biology as a biology-based inter-disciplinary study field that focuses on complex interactions in biological systems, claiming that it uses...
approaches such as interactomics
Interactomics
Interactomics is a discipline at the intersection of bioinformatics and biology that deals with studying both the interactions and the consequences of those interactions between and among proteins, and other molecules within a cell. The network of all such interactions is called the Interactome...
give clues to a proteins function based on what it interacts with.
Crystallins
Cataract
A cataract is a clouding that develops in the crystalline lens of the eye or in its envelope, varying in degree from slight to complete opacity and obstructing the passage of light...
. However, recent studies have shown that the lens crystallins are much more diverse than previously recognized and that many are related or identical to metabolic enzymes and stress proteins found in numerous tissues. Unlike other proteins performing highly specialized tasks, such as globin
Globin
Globins are a related family of proteins, which are thought to share a common ancestor. These proteins all incorporate the globin fold, a series of eight alpha helical segments. Two prominent members of this family include myoglobin and hemoglobin, which both bind the heme prosthetic group...
or rhodopsin
Rhodopsin
Rhodopsin, also known as visual purple, is a biological pigment of the retina that is responsible for both the formation of the photoreceptor cells and the first events in the perception of light. Rhodopsins belong to the G-protein coupled receptor family and are extremely sensitive to light,...
, the crystallins are very diverse and show numerous species differences. Essentially all vertebrate lenses contain representatives of the α and β/γ crystallins, the "Ubiquitous crystallins", which are themselves heterogeneous, and only few species or selected taxonomic groups use entirely different proteins as lens crystallins.This paradox of crystallins being highly conserved in sequence while extremely diverse in number and distribution shows that many crystallins have vital functions outside the lens and cornea, and this multi-functionality of the crystallins is achieved by gene sharing.
Gene regulation
Crystallin recruitment may occur by changes in gene regulation that leads to high lens expression. One such example is gluthathione S-transferase/S11-crystallin that was specialized for lens expression by change in gene regulation and gene duplicationGene duplication
Gene duplication is any duplication of a region of DNA that contains a gene; it may occur as an error in homologous recombination, a retrotransposition event, or duplication of an entire chromosome.The second copy of the gene is often free from selective pressure — that is, mutations of it have no...
. The fact that similar transcriptional factors such as Pax-6, and retinoic acid receptors, regulate different crystalline genes, suggests that lens-specific expression have played a crucial role for recruiting multifunctional protein as crystallins. Crystallin recruitment has occurred both with and without gene duplication, and tandem gene duplication has taken place among some of the crystallins with one of the duplicates specializing for lens expression. Ubiquitous α –crystallins and bird δ –crystallins are two examples.
Alpha crystallins
The α-crystallins, which contributed to the discovery of crystallins as borrowed proteins, have continually supported the theory of gene sharing, and helped delineating the mechanisms used for gene sharing as well. There are two α-crystallin genes (αA and αB), which are about 55% identical in amino acid sequence. Expression studies in non-lens cells showed that the αB-crystallin, other than being a functional lens protein, is a functional small heat shock protein. αB-crystallin is induced by heat and other physiological stresses, and it can protect the cells from elevated temperatures and hypertonic stress. αB-crystallin is also overexpressed in many pathologies, including neurodegenerative diseases, fibroblasts of patients with Werner's disease showing premature senescence, and growth abnormalities. In addition to being overexpressed under abnormal conditions, αB-crystallin is constitutively expressed in heart, skeletal muscle, kidney, lung and many other tissues. In contrast to αB-crystallin, except for low-level expression in the thymus, spleen and retina, αA-crystallin is highly specialized for expression in the lens and is not stress-inducible. However, like αB-crystallin, it can also function as molecular chaperone and protect against thermal stress.Beta/gamma-crystallins
β/γ-crystallins are different from α-crystallins in that they are a large multigene family. Other proteins like bacterial spore coat, a slime mold cyst protein, and epidermis differentiation-specific protein, contain the same Greek key motifs and are placed under β/γ crystallin superfamily. This relationship supports the idea that β/γ- crystallins have been recruited by a gene-sharing mechanism. However, except for few reports, non-refractive function of the β/γ-crystallin is yet to be found.Corneal crystallins
Similar to lensLens (anatomy)
The crystalline lens is a transparent, biconvex structure in the eye that, along with the cornea, helps to refract light to be focused on the retina. The lens, by changing shape, functions to change the focal distance of the eye so that it can focus on objects at various distances, thus allowing a...
, cornea
Cornea
The cornea is the transparent front part of the eye that covers the iris, pupil, and anterior chamber. Together with the lens, the cornea refracts light, with the cornea accounting for approximately two-thirds of the eye's total optical power. In humans, the refractive power of the cornea is...
is a transparent, avascular tissue derived from the ectoderm
Ectoderm
The "ectoderm" is one of the three primary germ cell layers in the very early embryo. The other two layers are the mesoderm and endoderm , with the ectoderm as the most exterior layer...
that is responsible for focusing light onto the retina
Retina
The vertebrate retina is a light-sensitive tissue lining the inner surface of the eye. The optics of the eye create an image of the visual world on the retina, which serves much the same function as the film in a camera. Light striking the retina initiates a cascade of chemical and electrical...
. However, unlike lens, cornea depends on the air-cell interface and its curvature for refraction. Early immunology studies have shown that BCP 54 comprises 20–40% of the total soluble protein in bovine cornea. Subsequent studies have indicated that BCP 54 is ALDH3, a tumor and xenobiotic-inducible cytosolic enzyme, found in human, rat, and other mammals.
Non refractive roles of crystallins in lens and cornea
While it is evident that gene sharing resulted in many of lens crystallins being multifunctional proteins, it is still uncertain to what extent the crystallins use their non-refractive properties in the lens, or on what basis they were selected. The α-crystallins provide a convincing case for a lens crystallin using its non-refractive ability within the lens to prevent protein aggregation under a variety of environmental stresses and to protect against enzyme inactivation by post-translational modifications such as glycationGlycation
Glycation is the result of the bonding of a protein or lipid molecule with a sugar molecule, such as fructose or glucose, without the controlling action of an enzyme. All blood sugars are reducing molecules. Glycation may occur either inside the body or outside the body...
. The α-crystallins may also play a functional role in the stability and remodeling of the cytoskeleton during fiber cell differentiation in the lens. In cornea, ALDH3 is also suggested to be responsible for absorbing UV-B light.
Co-evolution of lens and cornea through gene sharing
Based on the similarities between lens and cornea, such as abundant water-soluble enzymes, and being derived from ectoderm, the lens and cornea are thought to be co-evolved as a "refraction unit." Gene sharing would maximize light transmission and refraction to the retina by this refraction unit. Studies have shown that many water soluble enzymes/proteins expressed by cornea are identical to taxon-specific lens crystallins, such as ALDH1A1/ η-crystallin, α-enolase/τ-crystallin, and lactic dehydrogenase/ -crystallin. Also, the anuran corneal epithelium, which can transdifferentiate to regenerate the lens, abundantly expresses ubiquitous lens crystallins, α, β and γ, in addition to the taxon-specific crystallin α-enolase/τ-crystallin. Overall, the similarity in expression of these proteins in the cornea and lens, both in abundance and taxon-specificity, supports the idea of co-evolution of lens and cornea through gene sharing.Relationship to similar concepts
Gene sharing is related to, but distinct from, several concepts in genetics, evolution, and molecular biology. Gene sharing entails multiple effects from the same gene, but unlike pleiotropyPleiotropy
Pleiotropy occurs when one gene influences multiple phenotypic traits. Consequently, a mutation in a pleiotropic gene may have an effect on some or all traits simultaneously...
, it necessarily involves separate functions at the molecular level. A gene could exhibit pleiotropy when single enzyme function affects multiple phenotypic traits
Trait (biology)
A trait is a distinct variant of a phenotypic character of an organism that may be inherited, environmentally determined or be a combination of the two...
; mutations of a shared gene could potentially affect only a single trait. Gene duplication
Gene duplication
Gene duplication is any duplication of a region of DNA that contains a gene; it may occur as an error in homologous recombination, a retrotransposition event, or duplication of an entire chromosome.The second copy of the gene is often free from selective pressure — that is, mutations of it have no...
followed by differential mutation is another phenomenon thought to be a key element in the evolution of protein function, but in gene sharing, there is no divergence of gene sequence when proteins take on new functions; the single polypeptide takes on new roles while retaining old ones. Alternative splicing
Alternative splicing
Alternative splicing is a process by which the exons of the RNA produced by transcription of a gene are reconnected in multiple ways during RNA splicing...
can result in the production of multiple polypeptides (with multiple functions) from a single gene, but by definition, gene sharing involves multiple functions of a single polypeptide.
Clinical significance
The multiple roles of moonlighting proteins complicates the determination of phenotypePhenotype
A phenotype is an organism's observable characteristics or traits: such as its morphology, development, biochemical or physiological properties, behavior, and products of behavior...
from genotype
Genotype
The genotype is the genetic makeup of a cell, an organism, or an individual usually with reference to a specific character under consideration...
, hampering the study of inherited metabolic disorder
Inborn error of metabolism
Inborn errors of metabolism comprise a large class of genetic diseases involving disorders of metabolism. The majority are due to defects of single genes that code for enzymes that facilitate conversion of various substances into others...
s.
The complex phenotypes of several disorders are suspected to be caused by the involvement of moonlighting proteins. The protein GAPDH
Glyceraldehyde 3-phosphate dehydrogenase
Glyceraldehyde 3-phosphate dehydrogenase is an enzyme of ~37kDa that catalyzes the sixth step of glycolysis and thus serves to break down glucose for energy and carbon molecules...
has at least 11 documented functions, one of which includes apoptosis. Excessive apoptosis is involved in many neurodegenerative diseases, such as Huntington's
Huntington's disease
Huntington's disease, chorea, or disorder , is a neurodegenerative genetic disorder that affects muscle coordination and leads to cognitive decline and dementia. It typically becomes noticeable in middle age. HD is the most common genetic cause of abnormal involuntary writhing movements called chorea...
, Alzheimer's
Alzheimer's disease
Alzheimer's disease also known in medical literature as Alzheimer disease is the most common form of dementia. There is no cure for the disease, which worsens as it progresses, and eventually leads to death...
, and Parkinson's
Parkinson's disease
Parkinson's disease is a degenerative disorder of the central nervous system...
as well as in brain ischemia
Ischemia
In medicine, ischemia is a restriction in blood supply, generally due to factors in the blood vessels, with resultant damage or dysfunction of tissue. It may also be spelled ischaemia or ischæmia...
. In one case, GAPDH was found in the degenerated neurons of individuals who had Alzheimer's disease.
Although there is insufficient evidence for definite conclusions, there are well documented examples of moonlighting proteins that play a role in disease. One such disease is tuberculosis
Tuberculosis
Tuberculosis, MTB, or TB is a common, and in many cases lethal, infectious disease caused by various strains of mycobacteria, usually Mycobacterium tuberculosis. Tuberculosis usually attacks the lungs but can also affect other parts of the body...
. One moonlighting protein in the bacterium M. tuberculosis
Mycobacterium tuberculosis
Mycobacterium tuberculosis is a pathogenic bacterial species in the genus Mycobacterium and the causative agent of most cases of tuberculosis . First discovered in 1882 by Robert Koch, M...
has a function which counteracts the effects of antibiotics. Specifically, M. tuberculosis gains antibiotic resistance
Antibiotic resistance
Antibiotic resistance is a type of drug resistance where a microorganism is able to survive exposure to an antibiotic. While a spontaneous or induced genetic mutation in bacteria may confer resistance to antimicrobial drugs, genes that confer resistance can be transferred between bacteria in a...
against ciprofloxacin
Ciprofloxacin
Ciprofloxacin is a synthetic chemotherapeutic antibiotic of the fluoroquinolone drug class.It is a second-generation fluoroquinolone antibacterial. It kills bacteria by interfering with the enzymes that cause DNA to rewind after being copied, which stops synthesis of DNA and of...
from overexpression of Glutamate racemase
Glutamate racemase
In enzymology, glutamate racemase is an enzyme that catalyzes the chemical reactionHence, this enzyme RacE has one substrate, L-glutamate, and one product, D-glutamate....
in vivo
In vivo
In vivo is experimentation using a whole, living organism as opposed to a partial or dead organism, or an in vitro controlled environment. Animal testing and clinical trials are two forms of in vivo research...
.