L-ribulose-5-phosphate 4-epimerase
Encyclopedia
In enzymology, a L-ribulose-5-phosphate 4-epimerase is an enzyme
that catalyzes
the interconversion of Ribulose 5-phosphate
and Xylulose 5-phosphate
in the oxidative phase of the Pentose phosphate pathway
.
This enzyme has a molecular mass of 102 kDa
and is believed to be composed of four identical 25.5 kDa subunits. It belongs to the family of isomerase
s, specifically those racemases and epimerases acting on carbohydrate
s and derivatives. The systematic name of this enzyme class is L-ribulose-5-phosphate 4-epimerase. Other names in common use include phosphoribulose isomerase, ribulose phosphate 4-epimerase, L-ribulose-phosphate 4-epimerase, L-ribulose 5-phosphate 4-epimerase, AraD, and L-Ru5P. This enzyme participates in pentose and glucuronate interconversions and ascorbate and aldarate metabolism.
to Xylulose 5-phosphate
by aldol cleavage and subsequent aldol condensation
. The proposed mechanism involves the abstraction of the proton
from the hydroxyl
group on C-4, followed by cleavage of the bond between C-3 and C-4 to give a metal-stabilized acetone enediolate and a glycolaldehyde
phosphate fragment. The C-C bond of glycolaldehyde phosphate is then rotated 180°, and the C-C bond between C-3 and C-4 is regenerated to give inversion of stereochemistry
at C-4.
This mechanism is contested by a possible alternative dehydration
reaction scheme. The literature favors the aldol mechanism for two reasons, first, the aldol cleavage-condesation mechanism is analogous to the reaction catalyzed by L-fuculose-phosphate aldolase
which has high levels of sequence similarity with Ribulose 5-Phosphate 4-Epimerase. Second, the analysis of 13C
and deuterium
kinetic isotope effect
s points toward the aldol mechanism. It has been reported that there is little to no difference in the deuterium isotope
effects at C-3 and C-4, suggesting that these hydrogen bonds are not broken during epimerization. Changes in isotope effect at C-3 would be expected for the dehydration mechanism, because the breaking of the C-H bond is the rate-limiting step in this mechanism and substituting the C-3 hydrogen
with deuterium would significantly alter the rate. At the same time there are significant levels of 13C isotope effects, suggesting C-C bond breakage as expected with the aldol mechanism.
has a single domain consisting of a central β sheet flanked on either side by layers of α-helix. A central β-sheet is formed from nine β-strands (b1-b9) and is predominantly antiparallel except between strands b7 and b8. The eight α-helices of the structure form two layers on either side of the central β-sheet. The active site is identified by the position of the catalytic zinc
residue and is located at the interface between two adjacent subunits. Asp76, His95, His97, and His171act as the metal-binding resdues. A remarkable feature of the structure is that it shows a very close resemblance to that of L-fuculose-phosphate aldolase
. This is consistent with the notion that both enzymes belong to a superfamily of epimerases/aldolase
s that catalyze carbon-carbon bond cleavage reactions via a metal-stabilized enolate intermediate.
. This operon consists of eight genes araA-araH with the gene for Ribulose 5-phosphate 4-epimerase called araD. The arabinose system enables the take up the pentose
L-arabinose, and then the conversion of intracellular
arabinose
in three steps catalyzed by the products of the araB, araA, araD genes to D-xylulose-5-phosphate.
s differing greatly in the position of the phosphate (C-5 vs C-1).
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...
that catalyzes
Catalysis
Catalysis is the change in rate of a chemical reaction due to the participation of a substance called a catalyst. Unlike other reagents that participate in the chemical reaction, a catalyst is not consumed by the reaction itself. A catalyst may participate in multiple chemical transformations....
the interconversion of Ribulose 5-phosphate
Ribulose 5-phosphate
Ribulose 5-phosphate is one of the end-products of the pentose phosphate pathway. It is also an intermediate in the Calvin cycle.It is formed by phosphogluconate dehydrogenase, and it can be acted upon by phosphopentose isomerase and phosphopentose epimerase....
and Xylulose 5-phosphate
Xylulose 5-phosphate
D-Xylulose 5-phosphate is an intermediate in the Pentose Phosphate Pathway. It is a ketose sugar formed from ribulose-5-phosphate...
in the oxidative phase of the Pentose phosphate pathway
Pentose phosphate pathway
The pentose phosphate pathway is a process that generates NADPH and pentoses . There are two distinct phases in the pathway. The first is the oxidative phase, in which NADPH is generated, and the second is the non-oxidative synthesis of 5-carbon sugars...
.
- L-ribulose 5-phosphate
D-xylulose 5-phosphate
This enzyme has a molecular mass of 102 kDa
KDA
KDA may refer to:* Karachi Development Authority* Kongsberg Defence & Aerospace* Kotelawala Defence Academy* Kramer Design Associates* Lithium diisopropylamide, KDA is the potassium analogue of lithium diisopropylamideOr kDa may refer to:...
and is believed to be composed of four identical 25.5 kDa subunits. It belongs to the family of isomerase
Isomerase
In biochemistry, an isomerase is an enzyme that catalyzes the structural rearrangement of isomers. Isomerases thus catalyze reactions of the formwhere B is an isomer of A.-Nomenclature:...
s, specifically those racemases and epimerases acting on carbohydrate
Carbohydrate
A carbohydrate is an organic compound with the empirical formula ; that is, consists only of carbon, hydrogen, and oxygen, with a hydrogen:oxygen atom ratio of 2:1 . However, there are exceptions to this. One common example would be deoxyribose, a component of DNA, which has the empirical...
s and derivatives. The systematic name of this enzyme class is L-ribulose-5-phosphate 4-epimerase. Other names in common use include phosphoribulose isomerase, ribulose phosphate 4-epimerase, L-ribulose-phosphate 4-epimerase, L-ribulose 5-phosphate 4-epimerase, AraD, and L-Ru5P. This enzyme participates in pentose and glucuronate interconversions and ascorbate and aldarate metabolism.
Enzyme Mechanism
Ribulose 5-phosphate 4-epimerase catalyzes the epimerization of Ribulose 5-phosphateRibulose 5-phosphate
Ribulose 5-phosphate is one of the end-products of the pentose phosphate pathway. It is also an intermediate in the Calvin cycle.It is formed by phosphogluconate dehydrogenase, and it can be acted upon by phosphopentose isomerase and phosphopentose epimerase....
to Xylulose 5-phosphate
Xylulose 5-phosphate
D-Xylulose 5-phosphate is an intermediate in the Pentose Phosphate Pathway. It is a ketose sugar formed from ribulose-5-phosphate...
by aldol cleavage and subsequent aldol condensation
Aldol condensation
An aldol condensation is an organic reaction in which an enol or an enolate ion reacts with a carbonyl compound to form a β-hydroxyaldehyde or β-hydroxyketone, followed by a dehydration to give a conjugated enone....
. The proposed mechanism involves the abstraction of the proton
Proton
The proton is a subatomic particle with the symbol or and a positive electric charge of 1 elementary charge. One or more protons are present in the nucleus of each atom, along with neutrons. The number of protons in each atom is its atomic number....
from the hydroxyl
Hydroxyl
A hydroxyl is a chemical group containing an oxygen atom covalently bonded with a hydrogen atom. In inorganic chemistry, the hydroxyl group is known as the hydroxide ion, and scientists and reference works generally use these different terms though they refer to the same chemical structure in...
group on C-4, followed by cleavage of the bond between C-3 and C-4 to give a metal-stabilized acetone enediolate and a glycolaldehyde
Glycolaldehyde
Glycolaldehyde is the smallest possible molecule that contains both an aldehyde group and a hydroxyl group. It is the only possible diose, a 2-carbon monosaccharide, although a diose is not strictly a saccharide...
phosphate fragment. The C-C bond of glycolaldehyde phosphate is then rotated 180°, and the C-C bond between C-3 and C-4 is regenerated to give inversion of stereochemistry
Stereochemistry
Stereochemistry, a subdiscipline of chemistry, involves the study of the relative spatial arrangement of atoms within molecules. An important branch of stereochemistry is the study of chiral molecules....
at C-4.
This mechanism is contested by a possible alternative dehydration
Dehydration reaction
In chemistry and the biological sciences, a dehydration reaction is usually defined as a chemical reaction that involves the loss of water from the reacting molecule. Dehydration reactions are a subset of elimination reactions...
reaction scheme. The literature favors the aldol mechanism for two reasons, first, the aldol cleavage-condesation mechanism is analogous to the reaction catalyzed by L-fuculose-phosphate aldolase
L-fuculose-phosphate aldolase
In enzymology, a L-fuculose-phosphate aldolase is an enzyme that catalyzes the chemical reactionHence, this enzyme has one substrate, L-fuculose-1-phosphate, and two products, glycerone phosphate and -lactaldehyde....
which has high levels of sequence similarity with Ribulose 5-Phosphate 4-Epimerase. Second, the analysis of 13C
Carbon-13
Carbon-13 is a natural, stable isotope of carbon and one of the environmental isotopes. It makes up about 1.1% of all natural carbon on Earth.- Detection by mass spectrometry :...
and deuterium
Deuterium
Deuterium, also called heavy hydrogen, is one of two stable isotopes of hydrogen. It has a natural abundance in Earth's oceans of about one atom in of hydrogen . Deuterium accounts for approximately 0.0156% of all naturally occurring hydrogen in Earth's oceans, while the most common isotope ...
kinetic isotope effect
Kinetic isotope effect
The kinetic isotope effect is the ratio of reaction rates of two different isotopically labeled molecules in a chemical reaction. It is also called "isotope fractionation," although this term is somewhat broader in meaning...
s points toward the aldol mechanism. It has been reported that there is little to no difference in the deuterium isotope
Isotope
Isotopes are variants of atoms of a particular chemical element, which have differing numbers of neutrons. Atoms of a particular element by definition must contain the same number of protons but may have a distinct number of neutrons which differs from atom to atom, without changing the designation...
effects at C-3 and C-4, suggesting that these hydrogen bonds are not broken during epimerization. Changes in isotope effect at C-3 would be expected for the dehydration mechanism, because the breaking of the C-H bond is the rate-limiting step in this mechanism and substituting the C-3 hydrogen
Hydrogen
Hydrogen is the chemical element with atomic number 1. It is represented by the symbol H. With an average atomic weight of , hydrogen is the lightest and most abundant chemical element, constituting roughly 75% of the Universe's chemical elemental mass. Stars in the main sequence are mainly...
with deuterium would significantly alter the rate. At the same time there are significant levels of 13C isotope effects, suggesting C-C bond breakage as expected with the aldol mechanism.
Structure
The structure is homo-tetrameric and displays C4 symmetry. Each protein subunitProtein subunit
In structural biology, a protein subunit or subunit protein is a single protein molecule that assembles with other protein molecules to form a protein complex: a multimeric or oligomeric protein. Many naturally occurring proteins and enzymes are multimeric...
has a single domain consisting of a central β sheet flanked on either side by layers of α-helix. A central β-sheet is formed from nine β-strands (b1-b9) and is predominantly antiparallel except between strands b7 and b8. The eight α-helices of the structure form two layers on either side of the central β-sheet. The active site is identified by the position of the catalytic zinc
Zinc
Zinc , or spelter , is a metallic chemical element; it has the symbol Zn and atomic number 30. It is the first element in group 12 of the periodic table. Zinc is, in some respects, chemically similar to magnesium, because its ion is of similar size and its only common oxidation state is +2...
residue and is located at the interface between two adjacent subunits. Asp76, His95, His97, and His171act as the metal-binding resdues. A remarkable feature of the structure is that it shows a very close resemblance to that of L-fuculose-phosphate aldolase
L-fuculose-phosphate aldolase
In enzymology, a L-fuculose-phosphate aldolase is an enzyme that catalyzes the chemical reactionHence, this enzyme has one substrate, L-fuculose-1-phosphate, and two products, glycerone phosphate and -lactaldehyde....
. This is consistent with the notion that both enzymes belong to a superfamily of epimerases/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 that catalyze carbon-carbon bond cleavage reactions via a metal-stabilized enolate intermediate.
Biological Function
Ribulose 5-phosphate 4-epimerase is found on the well studied L-arabinose operonL-arabinose operon
The L-arabinose operon of the model bacterium Escherichia coli has been a focus for research in molecular biology for over 40 years, and has been investigated extensively at the genetic, biochemical, physiological, and biophysical levels....
. This operon consists of eight genes araA-araH with the gene for Ribulose 5-phosphate 4-epimerase called araD. The arabinose system enables the take up the pentose
Pentose
A pentose is a monosaccharide with five carbon atoms. Pentoses are organized into two groups. Aldopentoses have an aldehyde functional group at position 1...
L-arabinose, and then the conversion of intracellular
Intracellular
Not to be confused with intercellular, meaning "between cells".In cell biology, molecular biology and related fields, the word intracellular means "inside the cell".It is used in contrast to extracellular...
arabinose
Arabinose
Arabinose is an aldopentose – a monosaccharide containing five carbon atoms, and including an aldehyde functional group.For biosynthetic reasons, most saccharides are almost always more abundant in nature as the "D"-form, or structurally analogous to D-glyceraldehyde.For sugars, the D/L...
in three steps catalyzed by the products of the araB, araA, araD genes to D-xylulose-5-phosphate.
| Gene | Protein |
|---|---|
| AraA | Isomerase |
| AraB | Ribulokinase |
| AraC | Regulatory |
| AraD | Epimerase |
| AraE | Uptake |
| AraF | Uptake |
| AraG | Uptake |
| AraH | Uptake |
Evolution
L-Ribulose-5-phosphate 4-epimerase and L-fuculose-1-phosphate (L-Fuc1P) aldolase are evolutionarily related enzymes that display 26% sequence identity and a very high degree of structural similarity. They both employ a divalent cation in the stabilization of an enolate during catalysis, and both are able to deprotonate the C-4 hydroxyl group of a phosphoketose substrate. Despite these many similarities, subtle distinctions are present which allow the enzymes to catalyze two seemingly different reactions and to accommodate substrateSubstrate
Substrate may mean:*Substrate , Natural stone, masonry surface, ceramic and porcelain tiles*Substrate , the material used in the bottom of an aquarium*Substrate , the material used in the bottom of a vivarium or terrarium...
s differing greatly in the position of the phosphate (C-5 vs C-1).