DNA-deoxyinosine glycosylase

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DNA-deoxyinosine glycosylase
Identifiers
EC no. 3.2.2.15
CAS no. 68247-62-1
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DNA-deoxyinosine glycosylase (EC 3.2.2.15, DNA(hypoxanthine) glycohydrolase, deoxyribonucleic acid glycosylase, hypoxanthine-DNA glycosylase) is an enzyme with systematic name DNA-deoxyinosine deoxyribohydrolase. [1] This enzyme is involved in DNA damage repair and targets hypoxanthine bases.

Contents

DNA is constantly exposed to chemical reactions within its cellular environment, leading to undesired structural changes and compromising the integrity of genetic information. Common changes to bases include oxidation, alkylation, or the deamination of bases. Observed deamination in DNA bases are cytosine to uracil, guanine to xanthine and oxanine, or adenine to hypoxanthine. These single-base legions are removed through the base excision repair (BER) pathway, a mechanism initiated by DNA glycosylase to remove a mismatch or mutated base. [2] [3]

DNA glycosylases are ubiquitous, observed in species of all kingdoms of life, including archaea, eubacteria, eukaryotes, and large DNA viruses. They can be further subdivided into four specific categories: the uracil DNA glycosylases (UDGs), the helix-hairpin-helix (HhH) glycosylases, the 3-methyl-purine glycosylase (MPG), and the endonuclease VIII-like (NEIL) glycosylases. [4] However, DNA deoxyinosine glycosylase activity, the removal of a hypoxanthine base, is observed within the uracil DNA glycosylase superfamily. [5] [6] Currently, there are 6 families within the DNA Uracil Glycosylase superfamily, each classified based on their sequence homology, and biochemical and structural similarities. [7] Uracil and hypoxanthine activity is a feature that is particular to UDG families, namely family 3 SMUG1-like enzymes and more recently the newly identified family 6 enzymes, which exhibits only hypoxanthine activity specifically. [6]

Nomenclature

Reaction mechanism

DNA damage is caused by environmental and endogenous agents, that can create highly mutagenic lesions or compromise genomic stability. In order to preserve genomic sequence information, cells must counteract DNA damage through one of its five major pathways: base excision repair (BER), nucleotide excision repair (NER), mismatch repair (MMR), homologous recombination (HR), and non-homologous end joining (NHEJ). [9]

While hypoxanthine, or inosine when attached to a ribose, is a naturally-occurring base in human tRNA or in wobble base pairing, it is mistakenly incorporated into DNA either through the deamination of an adenine base pair, or as inosine triphosphate (ITP), formed through ATP deamination. [5] The mutated base gets removed through the base excision repair pathway, a two step mechanism carried out by a glycosylase and AP endonuclease. DNA deoxyinosine glycosylase initiates the process through hydrolytic cleavage of the hypoxanthine base, releasing a free hypoxanthine and creating an abasic, or AP site. AP endonuclease follows glycosylase activity by recognizing this AP site and nicking the phosphodiester backbone at the specified location. DNA polymerase and DNA ligase then completes the process by incorporating the correct base pair and closing the nick. [2]

PDB 4SKN model suggests catalytic mechanism of UDG family, which includes baseflipping mechanism in which uracil interacts with a uracil-binding pocket. PDB 4skn EBI.jpg
PDB 4SKN model suggests catalytic mechanism of UDG family, which includes baseflipping mechanism in which uracil interacts with a uracil-binding pocket.

The UDG superfamily typically contains a four-strand β-sheet surrounded by α-helices and undergoes the same or a similar mechanism to create abasic sites. The general catalytic mechanism involves activating the leaving group, stabilization of the oxocarbenium ion, and positioning of a water molecule. [10] Structural studies on human uracil DNA glycosylase (PDB 4KSN) suggests a base-flipping mechanism that requires a few key residues. L272 allows the enzyme to intercalate into the major groove of DNA, allowing additional interactions of uracil with the uracil-binding pocket, which in this model includes His 268, Gin 144, Phe 158, Asn 145, Asn 204, and Tyr 147. When uracil sits in the binding pocket, it is flipped out of the DNA base-stack and in proximity to the water molecule activated by residue D145. Hydrogen bonding between water and aspartate prepares the water molecule for nucleophilic attack of the N-glycosidic bond, cleaving a base and leaving behind an AP site. [11]

Structure

Recent crystal structure and phylogenetic studies of the family 3 SMUG1-like DNA glycoyslase from Pedobacter heparinus (PDB 5H0K) reveals both hypoxanthine/xanthine and uracil activity and has been classified into a new subfamily, Family 6. While crystal studies reveal folds that share similarity to SMUG1 enzymes, the phe SMUG2 UDG displays distinct differences in local structure based on a comparison of the three motifs in all UDG families that are involved in recognition and catalysis. Analysis of motif 1 reveals a GINPG sequence, similar to family 2 UDGs. The N63 residue is highly conserved and also appears in families 2 and 3, presumably key for the positioning of the water molecule. S124 is the first residue in motif 3 phe SMUG2, but is typically an asparagine in other UDG families. Curiously, a mutation in which serine is replaced with asparagine reveals increased catalytic activity as well as broadens activity to include single-stranded uracil-containing DNA and G/T base pairs. This suggests that S124 may play a key role in increasing substrate specificity. Motif 2 begins with highly conserved H205, which aids in UDG activity by forming short-distance hydrogen bonds with the O2 of uracil. While crucial for UDG activity involving A/U or G/U recognition, mutation of this residue has no effect on hypoxanthine and xanthine DNA glycosylase activity, however. [6]

Related Research Articles

<span class="mw-page-title-main">Hypoxanthine</span> Chemical compound

Hypoxanthine is a naturally occurring purine derivative. It is occasionally found as a constituent of nucleic acids, where it is present in the anticodon of tRNA in the form of its nucleoside inosine. It has a tautomer known as 6-hydroxypurine. Hypoxanthine is a necessary additive in certain cells, bacteria, and parasite cultures as a substrate and nitrogen source. For example, it is commonly a required reagent in malaria parasite cultures, since Plasmodium falciparum requires a source of hypoxanthine for nucleic acid synthesis and energy metabolism.

Deamination is the removal of an amino group from a molecule. Enzymes that catalyse this reaction are called deaminases.

<span class="mw-page-title-main">Nuclease</span> Class of enzymes which cleave nucleic acids

In biochemistry, a nuclease is an enzyme capable of cleaving the phosphodiester bonds between nucleotides of nucleic acids. Nucleases variously effect single and double stranded breaks in their target molecules. In living organisms, they are essential machinery for many aspects of DNA repair. Defects in certain nucleases can cause genetic instability or immunodeficiency. Nucleases are also extensively used in molecular cloning.

DNA glycosylases are a family of enzymes involved in base excision repair, classified under EC number EC 3.2.2. Base excision repair is the mechanism by which damaged bases in DNA are removed and replaced. DNA glycosylases catalyze the first step of this process. They remove the damaged nitrogenous base while leaving the sugar-phosphate backbone intact, creating an apurinic/apyrimidinic site, commonly referred to as an AP site. This is accomplished by flipping the damaged base out of the double helix followed by cleavage of the N-glycosidic bond.

<span class="mw-page-title-main">AP site</span> Biochemical site of damaged DNA or RNA

In biochemistry and molecular genetics, an AP site, also known as an abasic site, is a location in DNA that has neither a purine nor a pyrimidine base, either spontaneously or due to DNA damage. It has been estimated that under physiological conditions 10,000 apurinic sites and 500 apyrimidinic may be generated in a cell daily.

<span class="mw-page-title-main">Base excision repair</span> DNA repair process

Base excision repair (BER) is a cellular mechanism, studied in the fields of biochemistry and genetics, that repairs damaged DNA throughout the cell cycle. It is responsible primarily for removing small, non-helix-distorting base lesions from the genome. The related nucleotide excision repair pathway repairs bulky helix-distorting lesions. BER is important for removing damaged bases that could otherwise cause mutations by mispairing or lead to breaks in DNA during replication. BER is initiated by DNA glycosylases, which recognize and remove specific damaged or inappropriate bases, forming AP sites. These are then cleaved by an AP endonuclease. The resulting single-strand break can then be processed by either short-patch or long-patch BER.

<span class="mw-page-title-main">AP endonuclease</span> Enzyme involved in DNA repair

Apurinic/apyrimidinic (AP) endonuclease is an enzyme that is involved in the DNA base excision repair pathway (BER). Its main role in the repair of damaged or mismatched nucleotides in DNA is to create a nick in the phosphodiester backbone of the AP site created when DNA glycosylase removes the damaged base.

The enzyme DNA-(apurinic or apyrimidinic site) lyase, also referred to as DNA-(apurinic or apyrimidinic site) 5'-phosphomonoester-lyase or DNA AP lyase catalyzes the cleavage of the C-O-P bond 3' from the apurinic or apyrimidinic site in DNA via β-elimination reaction, leaving a 3'-terminal unsaturated sugar and a product with a terminal 5'-phosphate. In the 1970s, this class of enzyme was found to repair at apurinic or apyrimidinic DNA sites in E. coli and in mammalian cells. The major active enzyme of this class in bacteria, and specifically, E. coli is endonuclease type III. This enzyme is part of a family of lyases that cleave carbon-oxygen bonds.

<span class="mw-page-title-main">Uracil-DNA glycosylase</span> Enzyme that repairs DNA damage

Uracil-DNA glycosylase is an enzyme. Its most important function is to prevent mutagenesis by eliminating uracil from DNA molecules by cleaving the N-glycosidic bond and initiating the base-excision repair (BER) pathway.

<span class="mw-page-title-main">Cyclin O</span> Protein-coding gene in the species Homo sapiens

Cyclin-O is a protein that in humans is encoded by the CCNO gene.

<span class="mw-page-title-main">DNA-3-methyladenine glycosylase</span> Protein-coding gene in the species Homo sapiens

DNA-3-methyladenine glycosylase also known as 3-alkyladenine DNA glycosylase (AAG) or N-methylpurine DNA glycosylase (MPG) is an enzyme that in humans is encoded by the MPG gene.

<span class="mw-page-title-main">NTHL1</span> Protein-coding gene in the species Homo sapiens

Endonuclease III-like protein 1 is an enzyme that in humans is encoded by the NTHL1 gene.

<span class="mw-page-title-main">NEIL1</span> Protein-coding gene in the species Homo sapiens

Endonuclease VIII-like 1 is an enzyme that in humans is encoded by the NEIL1 gene.

<span class="mw-page-title-main">MBD4</span> Protein-coding gene in the species Homo sapiens

Methyl-CpG-binding domain protein 4 is a protein that in humans is encoded by the MBD4 gene.

<span class="mw-page-title-main">SMUG1</span> Protein-coding gene in the species Homo sapiens

Single-strand selective monofunctional uracil DNA glycosylase is an enzyme that in humans is encoded by the SMUG1 gene. SMUG1 is a glycosylase that removes uracil from single- and double-stranded DNA in nuclear chromatin, thus contributing to base excision repair.

<span class="mw-page-title-main">FPG IleRS zinc finger</span>

The FPG IleRS zinc finger domain represents a zinc finger domain found at the C-terminal in both DNA glycosylase/AP lyase enzymes and in isoleucyl tRNA synthetase. In these two types of enzymes, the C-terminal domain forms a zinc finger.

<span class="mw-page-title-main">H2TH domain</span>

In molecular biology, the H2TH domain is a DNA-binding domain found in DNA glycosylase/AP lyase enzymes, which are involved in base excision repair of DNA damaged by oxidation or by mutagenic agents. Most damage to bases in DNA is repaired by the base excision repair pathway. These enzymes are primarily from bacteria, and have both DNA glycosylase activity EC 3.2.2.- and AP lyase activity EC 4.2.99.18. Examples include formamidopyrimidine-DNA glycosylases and endonuclease VIII (Nei).

Deoxyribodipyrimidine endonucleosidase is an enzyme with systematic name deoxy-D-ribocyclobutadipyrimidine polynucleotidodeoxyribohydrolase. This enzyme catalyses the following chemical reaction

Double-stranded uracil-DNA glycosylase is an enzyme with systematic name uracil-double-stranded DNA deoxyribohydrolase (uracil-releasing). This enzyme catalyses a specific chemical reaction: it hydrolyses mismatched double-stranded DNA and polynucleotides, releasing free uracil.

<span class="mw-page-title-main">DNA base flipping</span> Biochemical process

DNA base flipping, or nucleotide flipping, is a mechanism in which a single nucleotide base, or nucleobase, is rotated outside the nucleic acid double helix. This occurs when a nucleic acid-processing enzyme needs access to the base to perform work on it, such as its excision for replacement with another base during DNA repair. It was first observed in 1994 using X-ray crystallography in a methyltransferase enzyme catalyzing methylation of a cytosine base in DNA. Since then, it has been shown to be used by different enzymes in many biological processes such as DNA methylation, various DNA repair mechanisms, and DNA replication. It can also occur in RNA double helices or in the DNA:RNA intermediates formed during RNA transcription.

References

  1. ↑ Karran P, Lindahl T (September 1978). "Enzymatic excision of free hypoxanthine from polydeoxynucleotides and DNA containing deoxyinosine monophosphate residues". The Journal of Biological Chemistry. 253 (17): 5877–5879. doi: 10.1016/S0021-9258(17)34545-3 . PMID   98523.
  2. 1 2 Lindahl T (1979-01-01). Cohn WE (ed.). "DNA glycosylases, endonucleases for apurinic/apyrimidinic sites, and base excision-repair". Progress in Nucleic Acid Research and Molecular Biology. Academic Press. 22: 135–192. doi:10.1016/s0079-6603(08)60800-4. ISBN   9780125400220. PMID   392601.
  3. ↑ Lindahl T, Wood RD (December 1999). "Quality control by DNA repair". Science. 286 (5446): 1897–1905. doi:10.1126/science.286.5446.1897. PMID   10583946.
  4. ↑ Jacobs AL, Schär P (February 2012). "DNA glycosylases: in DNA repair and beyond". Chromosoma. 121 (1): 1–20. doi:10.1007/s00412-011-0347-4. PMC   3260424 . PMID   22048164.
  5. 1 2 Lin T, Zhang L, Wu M, Jiang D, Li Z, Yang Z (2021). "Repair of Hypoxanthine in DNA Revealed by DNA Glycosylases and Endonucleases From Hyperthermophilic Archaea". Frontiers in Microbiology. 12: 736915. doi: 10.3389/fmicb.2021.736915 . PMC   8438529 . PMID   34531846.
  6. 1 2 3 Pang P, Yang Y, Li J, Wang Z, Cao W, Xie W (March 2017). "SMUG2 DNA glycosylase from Pedobacter heparinus as a new subfamily of the UDG superfamily". The Biochemical Journal. 474 (6): 923–938. doi: 10.1042/BCJ20160934 . PMID   28049757.
  7. ↑ Pearl LH (August 2000). "Structure and function in the uracil-DNA glycosylase superfamily". Mutation Research. 460 (3–4): 165–181. doi:10.1016/S0921-8777(00)00025-2. PMID   10946227.
  8. ↑ McDonald AG, Boyce S, Tipton KF (2001-05-30). "Enzyme classification and nomenclature". eLS (1st ed.). John Wiley & Sons, Ltd. pp. 1–11. doi:10.1002/9780470015902.a0000710.pub3. ISBN   978-0-470-01617-6.
  9. ↑ Chatterjee N, Walker GC (June 2017). "Mechanisms of DNA damage, repair, and mutagenesis". Environmental and Molecular Mutagenesis. 58 (5): 235–263. doi:10.1002/em.22087. PMC   5474181 . PMID   28485537.
  10. ↑ Lee HW, Dominy BN, Cao W (September 2011). "New family of deamination repair enzymes in uracil-DNA glycosylase superfamily". The Journal of Biological Chemistry. 286 (36): 31282–31287. doi: 10.1074/jbc.M111.249524 . PMC   3173141 . PMID   21642431.
  11. ↑ Slupphaug G, Mol CD, Kavli B, Arvai AS, Krokan HE, Tainer JA (November 1996). "A nucleotide-flipping mechanism from the structure of human uracil-DNA glycosylase bound to DNA". Nature. 384 (6604): 87–92. Bibcode:1996Natur.384...87S. doi:10.1038/384087a0. PMID   8900285. S2CID   4310250.
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