APOBEC3G
Encyclopedia
APOBEC3G is a human enzyme
encoded by the APOBEC3G gene
that belongs to the APOBEC
superfamily of protein
s. This family of proteins has been suggested to play an important role in innate anti-viral immunity
. APOBEC3G is a cytadine deaminase enzyme and is therefore known as DNA dC->dU-editing enzyme APOBEC-3G and APOBEC-related cytidine deaminase (ARCD).
APOBEC3G exerts innate antiretroviral immune activity against retrovirus
es, most notably HIV
, by interfering with proper replication. However, lentiviruses such as HIV have evolved the Viral infectivity factor
(Vif) protein in order to counteract this effect. Vif interacts with APOBEC3G and triggers the ubiquitination and degradation of APOBEC3G via the proteasomal pathway.
. Soon after, it was shown that APOBEC3G belonged to a family of proteins grouped together due to their homology with the cytidine deaminase APOBEC1.
between two beta sheet
s in the catalytic domain that could be a cofactor
binding site. (Figure 1)
CD2 is catalytically active and vital for deamination and motif specificity. CD1 is catalytically inactive, but very important for binding to DNA
and RNA
and is key to defining the 5’->3’ processivity of APOBEC3G deamination. Interestingly, CD2 is unable to exert deaminase activity without the presence of CD1.
“Native [APOBEC3G] is composed of monomer
s, dimer
s, trimers, tetramers, and higher order oligomer
s.” While it is thought that APOBEC3G functions as a dimer, it is possible that it actually functions as a mix of monomers and oligomers.
The D128 amino acid
residue, which lies within CD1 (Figure 1), appears to be particularly important for APOBEC3G interactions with Vif because a D128K point mutation prevents Vif-dependent depletion of APOBEC3G. Additionally, amino acids 128-130 on APOBEC3G form a negatively charged motif that is critical for interactions with Vif and the formation of APOBEC3G-Vif complexes. Furthermore, residues 124-127 are important for encapsidation of APOBEC3G into HIV-1 virions and the resulting antiretroviral activity.
s (AID). APOBEC3G interferes with reverse transcription by inducing numerous deoxycytidine
to deoxyuridine
mutations in the negative strand of the HIV DNA primarily expressed as complementary DNA
(cDNA) in a 3’->5’processive manner. Because APOBEC3G is part of the APOBEC superfamily and acts as an AID, it is likely that the mechanism mediated by APOBEC3G for cytidine deamination is similar to that of an E. coli cytidine deaminase that is known to be highly homologous to APOBEC1 and AID around the nucleotide and zinc binding region. The predicted deamination reaction is driven by a direct nucleophilic attack on position 4 of the cytidine pyrimidine
ring by the zinc-coordinated enzyme. Water is needed as both a hydrogen ion and hydroxyl
group donor (Figure 2). The deamination (and resulting oxidation) at position 4 yields a carbonyl group and results in a change from cytidine to uridine.
The deamination activity ultimately results in G→A hypermutations at “hot spots” of the proviral DNA. Such hypermutation ultimately destroys the coding and replicative capacity of the virus, resulting in many nonviable virions. APOBEC3G has a much weaker antiviral effect when its active site has been mutated to the point that the protein can no longer mutate retroviral DNA. The APOBEC3G-mediated deamination can also indirectly lead to viral DNA degradation by DNA repair systems attracted to the mutated residues.
to initiate reverse transcription. APOBEC3G can inhibit tRNA3Lys priming, thereby negatively affecting viral ssDNA production and virus infectivity. It is predicted that reverse transcription is also negatively affected by APOBEC3G binding to viral RNA and causing steric alterations.
(LTR) DNA ends.” These viral DNA ends “are inefficient substrates for integration and plus-strand DNA transfer.” As a result, HIV-1 provirus formation is inhibited.
T lymphocytes and macrophages. The encapsidation of APOBEC3G into HIV-1 virions is very important for the spread of APOBEC3G and the exertion of anti-retroviral activity. Encapsidation of APOBEC3G may occur by at least the following four proposed mechanisms (Figure 3): 1. Non-specific packaging of APOBEC3G 2. APOBEC3G interaction with host RNA 3. APOBEC3G interaction with viral RNA 4. Interaction of APOBEC3G with HIV-1 Gag proteins. Only the latter two mechanisms have been extensively confirmed.
The amount that is incorporated into virions is dependent on the level of APOBEC3G expression within the cell producing the virion. Xu et al. conducted studies with PBMC
cells and found that, in the absence of Vif, 7±4 APOBEC3G molecules were incorporated into the virions and resulted in potent inhibition of HIV-1 replication.
While APOBEC3G has typically been studied as a vital protein exhibiting potent antiviral effects on HIV-1, recent studies have elucidated the potential of APOBEC3G-mediated mutation to help to facilitate the propagation HIV-1. The number of deaminations in the preferred regions varies from one to many, possibly dependent on the time of exposure to APOBEC3G. Additionally, it has been shown that there is a dose response between intracellular APOBEC3G concentration and degree of viral hypermutation. Some HIV-1 provirus
es with APOBEC3G-mediated mutation have been shown to thrive because they carry too few mutations at APOBEC3G hotspots or because recombination between a lethally APOBEC3G-restricted provirus and a viable provirus has occurred. Such sublethal mutagenesis contributes to greater genetic diversity
among the HIV-1 virus population, demonstrating the potential for APOBEC3G to enhance HIV-1’s ability to adapt and propagate.
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...
encoded by the APOBEC3G gene
Gene
A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...
that belongs to the APOBEC
APOBEC
300px|thumb|upright|alt = Colored dice with checkered background|Example of a member of the APOBEC family, APOBEC-2. A cytidine deaminase from Homo Sapiens....
superfamily of protein
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...
s. This family of proteins has been suggested to play an important role in innate anti-viral immunity
Immunity (medical)
Immunity is a biological term that describes a state of having sufficient biological defenses to avoid infection, disease, or other unwanted biological invasion. Immunity involves both specific and non-specific components. The non-specific components act either as barriers or as eliminators of wide...
. APOBEC3G is a cytadine deaminase enzyme and is therefore known as DNA dC->dU-editing enzyme APOBEC-3G and APOBEC-related cytidine deaminase (ARCD).
APOBEC3G exerts innate antiretroviral immune activity against retrovirus
Retrovirus
A retrovirus is an RNA virus that is duplicated in a host cell using the reverse transcriptase enzyme to produce DNA from its RNA genome. The DNA is then incorporated into the host's genome by an integrase enzyme. The virus thereafter replicates as part of the host cell's DNA...
es, most notably HIV
HIV
Human immunodeficiency virus is a lentivirus that causes acquired immunodeficiency syndrome , a condition in humans in which progressive failure of the immune system allows life-threatening opportunistic infections and cancers to thrive...
, by interfering with proper replication. However, lentiviruses such as HIV have evolved the Viral infectivity factor
Viral infectivity factor
Viral infectivity factor, or Vif, is a protein found in HIV and other retroviruses. Its role is to disrupt the antiviral activity of the human enzyme APOBEC by targeting it for ubiquitination and cellular degradation. APOBEC is a cytidine deaminase enzyme that mutates viral nucleic acids.Vif is a...
(Vif) protein in order to counteract this effect. Vif interacts with APOBEC3G and triggers the ubiquitination and degradation of APOBEC3G via the proteasomal pathway.
Discovery
It was first identified by Jarmuz et al. as a member of family of proteins APOBEC3A to 3G on chromosome 22 in 2002 and later also as a cellular factor able to restrict replication of HIV-1 lacking the viral accessory protein VifViral infectivity factor
Viral infectivity factor, or Vif, is a protein found in HIV and other retroviruses. Its role is to disrupt the antiviral activity of the human enzyme APOBEC by targeting it for ubiquitination and cellular degradation. APOBEC is a cytidine deaminase enzyme that mutates viral nucleic acids.Vif is a...
. Soon after, it was shown that APOBEC3G belonged to a family of proteins grouped together due to their homology with the cytidine deaminase APOBEC1.
Structure
APOBEC3G has a symmetric structure, resulting in 2 homologous catalytic domains, the N-terminal (CD1) and C-terminal (CD2) domains, each of which contains a Zn2+ coordination site. Each domain also has the typical His/Cys-X-Glu-X23–28-Pro-Cys-X2-Cys motif for cytidine deaminases. However, unlike the typical cytidine deaminases, APOBEC3G contains a unique alpha helixAlpha helix
A common motif in the secondary structure of proteins, the alpha helix is a right-handed coiled or spiral conformation, in which every backbone N-H group donates a hydrogen bond to the backbone C=O group of the amino acid four residues earlier...
between two beta sheet
Beta sheet
The β sheet is the second form of regular secondary structure in proteins, only somewhat less common than the alpha helix. Beta sheets consist of beta strands connected laterally by at least two or three backbone hydrogen bonds, forming a generally twisted, pleated sheet...
s in the catalytic domain that could be a cofactor
Cofactor
Cofactor may refer to any of the following:* Cofactor , the signed minor of a matrix* Minor , an alternative name for the determinant of a smaller matrix than that which it describes...
binding site. (Figure 1)
CD2 is catalytically active and vital for deamination and motif specificity. CD1 is catalytically inactive, but very important for binding to DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
and RNA
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....
and is key to defining the 5’->3’ processivity of APOBEC3G deamination. Interestingly, CD2 is unable to exert deaminase activity without the presence of CD1.
“Native [APOBEC3G] is composed of monomer
Monomer
A monomer is an atom or a small molecule that may bind chemically to other monomers to form a polymer; the term "monomeric protein" may also be used to describe one of the proteins making up a multiprotein complex...
s, dimer
Protein dimer
In biochemistry, a dimer is a macromolecular complex formed by two, usually non-covalently bound, macromolecules like proteins or nucleic acids...
s, trimers, tetramers, and higher order oligomer
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...
s.” While it is thought that APOBEC3G functions as a dimer, it is possible that it actually functions as a mix of monomers and oligomers.
The D128 amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
residue, which lies within CD1 (Figure 1), appears to be particularly important for APOBEC3G interactions with Vif because a D128K point mutation prevents Vif-dependent depletion of APOBEC3G. Additionally, amino acids 128-130 on APOBEC3G form a negatively charged motif that is critical for interactions with Vif and the formation of APOBEC3G-Vif complexes. Furthermore, residues 124-127 are important for encapsidation of APOBEC3G into HIV-1 virions and the resulting antiretroviral activity.
Mechanism of Action
APOBEC3G has been widely studied and several mechanisms that negatively affect HIV-1 replication have been identified.Cytidine Deamination and Hypermutation
APOBEC3G and other proteins in the same family are able to act as Activation-Induced (Cytidine) DeaminaseActivation-Induced (Cytidine) Deaminase
Activation-induced deaminase is a 24 kDa enzyme that creates deliberate mutations in DNA.AID removes the amino group from a cytidine base, turning it into a uridine...
s (AID). APOBEC3G interferes with reverse transcription by inducing numerous deoxycytidine
Deoxycytidine
Deoxycytidine is a deoxyribonucleoside. It is like cytidine, but with one oxygen atom removed....
to deoxyuridine
Deoxyuridine
Deoxyuridine is a compound and a nucleoside. It is similar in chemical structure to uridine, but without the 2'-hydroxyl group.Idoxuridine and Trifluridine are variants of deoxyuridine used as antiviral drugs...
mutations in the negative strand of the HIV DNA primarily expressed as complementary DNA
Complementary DNA
In genetics, complementary DNA is DNA synthesized from a messenger RNA template in a reaction catalyzed by the enzyme reverse transcriptase and the enzyme DNA polymerase. cDNA is often used to clone eukaryotic genes in prokaryotes...
(cDNA) in a 3’->5’processive manner. Because APOBEC3G is part of the APOBEC superfamily and acts as an AID, it is likely that the mechanism mediated by APOBEC3G for cytidine deamination is similar to that of an E. coli cytidine deaminase that is known to be highly homologous to APOBEC1 and AID around the nucleotide and zinc binding region. The predicted deamination reaction is driven by a direct nucleophilic attack on position 4 of the cytidine pyrimidine
Pyrimidine
Pyrimidine is a heterocyclic aromatic organic compound similar to benzene and pyridine, containing two nitrogen atoms at positions 1 and 3 of the six-member ring...
ring by the zinc-coordinated enzyme. Water is needed as both a hydrogen ion and 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 donor (Figure 2). The deamination (and resulting oxidation) at position 4 yields a carbonyl group and results in a change from cytidine to uridine.
The deamination activity ultimately results in G→A hypermutations at “hot spots” of the proviral DNA. Such hypermutation ultimately destroys the coding and replicative capacity of the virus, resulting in many nonviable virions. APOBEC3G has a much weaker antiviral effect when its active site has been mutated to the point that the protein can no longer mutate retroviral DNA. The APOBEC3G-mediated deamination can also indirectly lead to viral DNA degradation by DNA repair systems attracted to the mutated residues.
Interference with Reverse Transcription
APOBEC3G interferes with reverse transcription of HIV-1 independent of DNA deamination. tRNA3Lys typically binds to the HIV-1 primer binding sitePrimer binding site
A primer binding site is a region of a nucleotide sequence where an RNA or DNA single-stranded primer binds to start replication. The primer binding site is on one of the two complementary strands of a double-stranded nucleotide polymer, in the strand which is to be copied, or is within a...
to initiate reverse transcription. APOBEC3G can inhibit tRNA3Lys priming, thereby negatively affecting viral ssDNA production and virus infectivity. It is predicted that reverse transcription is also negatively affected by APOBEC3G binding to viral RNA and causing steric alterations.
Interference with viral DNA integration
APOBEC3G was associated with interference of viral DNA integration into the host genome in a manner dependent on functional catalytic domains and deaminase activity. Mbisa et al. saw that APOBEC3G interferes with the processing and removal of primer tRNA from the DNA minus strand, thus leading to “aberrant viral 3’ long terminal repeatLong terminal repeat
Long terminal repeats are sequences of DNA that repeat hundreds or thousands of times. They are found in retroviral DNA and in retrotransposons, flanking functional genes...
(LTR) DNA ends.” These viral DNA ends “are inefficient substrates for integration and plus-strand DNA transfer.” As a result, HIV-1 provirus formation is inhibited.
Biological Function
APOBEC3G mRNA is expressed in certain cells, referred to as non-permissive cells, in which HIV-1 cannot properly infect and replicate in the absence of Vif. Such cells include physiologically relevant primary CD4CD4
CD4 is a glycoprotein expressed on the surface of T helper cells, monocytes, macrophages, and dendritic cells. It was discovered in the late 1970s and was originally known as leu-3 and T4 before being named CD4 in 1984...
T lymphocytes and macrophages. The encapsidation of APOBEC3G into HIV-1 virions is very important for the spread of APOBEC3G and the exertion of anti-retroviral activity. Encapsidation of APOBEC3G may occur by at least the following four proposed mechanisms (Figure 3): 1. Non-specific packaging of APOBEC3G 2. APOBEC3G interaction with host RNA 3. APOBEC3G interaction with viral RNA 4. Interaction of APOBEC3G with HIV-1 Gag proteins. Only the latter two mechanisms have been extensively confirmed.
The amount that is incorporated into virions is dependent on the level of APOBEC3G expression within the cell producing the virion. Xu et al. conducted studies with PBMC
PBMC
A peripheral blood mononuclear cell is any blood cell having a round nucleus. For example: a lymphocyte, a monocyte or a macrophage. These blood cells are a critical component in the immune system to fight infection and adapt to intruders. The lymphocyte population consists of T cells , B cells...
cells and found that, in the absence of Vif, 7±4 APOBEC3G molecules were incorporated into the virions and resulted in potent inhibition of HIV-1 replication.
Disease Relevance
APOBEC3G is expressed within the non-permissive cells and is a key inhibitory factor of HIV-1 replication and infectivity. However, Vif counteracts this antiretroviral factor, enabling production of viable and infective HIV-1 virions in the presence of APOBEC3G activity . In particular, Vif prevents incorporation of APOBEC3G into HIV-1 virions and promotes destruction of the enzyme in a manner independent of all other HIV-1 proteins.While APOBEC3G has typically been studied as a vital protein exhibiting potent antiviral effects on HIV-1, recent studies have elucidated the potential of APOBEC3G-mediated mutation to help to facilitate the propagation HIV-1. The number of deaminations in the preferred regions varies from one to many, possibly dependent on the time of exposure to APOBEC3G. Additionally, it has been shown that there is a dose response between intracellular APOBEC3G concentration and degree of viral hypermutation. Some HIV-1 provirus
Provirus
A provirus is a virus genome that is integrated into the DNA of a host cell.This state can be a stage of virus replication, or a state that persists over longer periods of time as either inactive viral infections or an endogenous retrovirus. In inactive viral infections the virus will not replicate...
es with APOBEC3G-mediated mutation have been shown to thrive because they carry too few mutations at APOBEC3G hotspots or because recombination between a lethally APOBEC3G-restricted provirus and a viable provirus has occurred. Such sublethal mutagenesis contributes to greater genetic diversity
Genetic diversity
Genetic diversity, the level of biodiversity, refers to the total number of genetic characteristics in the genetic makeup of a species. It is distinguished from genetic variability, which describes the tendency of genetic characteristics to vary....
among the HIV-1 virus population, demonstrating the potential for APOBEC3G to enhance HIV-1’s ability to adapt and propagate.