AlkB

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AlkB (Alkylation B) is a protein found in E. coli, induced during an adaptive response and involved in the direct reversal of alkylation damage. [1] [2] AlkB specifically removes alkylation damage to single stranded (SS) DNA caused by SN2 type of chemical agents. [3] It efficiently removes methyl groups from 1-methyl adenines, 3-methyl cytosines in SS DNA. [1] [4] AlkB is an alpha-ketoglutarate-dependent hydroxylase, a superfamily non-haem iron-containing proteins. It oxidatively demethylates the DNA substrate. [1] [4] Demethylation by AlkB is accompanied with release of CO2, succinate, and formaldehyde. [4]

Contents

Human homologs

There are nine human homologs of AlkB. [1] They are:

ABH3, like E. coli AlkB, is specific for SS DNA and RNA [1] whereas ABH2 has higher affinity for damages in double-stranded DNA. [5]

ALKBH8 has a RNA recognition motif, a methyltransferase domain, and an AlkB-like domain. The methyltransferase domain generates the wobble nucleoside 5-methoxycarbonylmethyluridine (mcm5U) from its precursor 5-carboxymethyluridine (cm5U). The AlkB-like domain generates (S)-5-methoxycarbonylhydroxymethyluridine (mchm5U)in Gly-tRNA-UCC. [6] [7]

FTO, which is associated with obesity in humans, is the first identified RNA demethylase. It demethylates N6-methyladenosine in mRNA. [8]

There is also another very different protein called AlkB or alkane hydroxylase. It is the catalytic subunit of a non-heme diiron protein, catalyzing the hydroxylation of alkanes, in aerobic bacteria that are able to utilize alkanes as a carbon source.

Virus homologs

AlkB domains are present within viral replication-associated proteins of plant RNA viruses of the families Closteroviridae, Alphaflexiviridae, Betaflexiviridae, and Secoviridae. [9] Potyviridae is the largest family of plant RNA viruses; [10] among these the AlkB domain is embedded in P1 proteases of endive necrotic mosaic virus (ENMV) of genus Potyvirus , French endive necrotic mosaic virus (FENMV) of Potyvirus, and blackberry virus Y (BlVY) of Brambyvirus. [11] [12]

Functions

AlkB has since been shown to have an ever expanding range of substrates since its initial discovery by Sedgwick, Lindahl, Seeberg and Falnes. Not only does it remove alkylation damage from the positively charged 1-methyl adenines and 3-methyl cytosines, but also from the neutral bases of 1-methyl guanine and 3-methyl thymine. [13] AlkB has been shown as the first example of a DNA repair enzyme converting one type of DNA damage that blocks DNA replication, to another type of damage that the DNA polymerase can traverse with ease. This was seen for the cyclic lesion ethanoadenine (not to be confused with ethenoadenine...see below), which upon hydroxylation by AlkB, affords an N6-acetaldehyde lesion, thus affording an 'adenine' hydrogen-bonding face. [14] In contrast to the previous types of alkylation damage removed by AlkB via a hydroxylation mechanism, AlkB has been shown to epoxidize the double bond of ethenoadenine, which is hydrolyzed to a diol, and ultimately released as the dialdehyde glyoxal, thus restoring the undamaged adenine in the DNA. [15]

Experimental results show that AlkB domains from plant viruses have RNA demethylase activity in vitro. [16] AlkB homologs from plants show the pro-viral roles, and may participate in plant antiviral immunity by regulating the levels of N6-methyladenosine (m6A), a common type of RNA modification. [11] [17] [18]

Related Research Articles

Methylation, in the chemical sciences, is the addition of a methyl group on a substrate, or the substitution of an atom by a methyl group. Methylation is a form of alkylation, with a methyl group replacing a hydrogen atom. These terms are commonly used in chemistry, biochemistry, soil science, and biology.

<span class="mw-page-title-main">DNA methyltransferase</span> Class of enzymes

In biochemistry, the DNA methyltransferase family of enzymes catalyze the transfer of a methyl group to DNA. DNA methylation serves a wide variety of biological functions. All the known DNA methyltransferases use S-adenosyl methionine (SAM) as the methyl donor.

<i>Potyviridae</i> Family of viruses

Potyviridae is a family of positive-strand RNA viruses that encompasses more than 30% of known plant viruses, many of which are of great agricultural significance. The family has 12 genera and 235 species, three of which are unassigned to a genus.

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">Methyltransferase</span> Group of methylating enzymes

Methyltransferases are a large group of enzymes that all methylate their substrates but can be split into several subclasses based on their structural features. The most common class of methyltransferases is class I, all of which contain a Rossmann fold for binding S-Adenosyl methionine (SAM). Class II methyltransferases contain a SET domain, which are exemplified by SET domain histone methyltransferases, and class III methyltransferases, which are membrane associated. Methyltransferases can also be grouped as different types utilizing different substrates in methyl transfer reactions. These types include protein methyltransferases, DNA/RNA methyltransferases, natural product methyltransferases, and non-SAM dependent methyltransferases. SAM is the classical methyl donor for methyltransferases, however, examples of other methyl donors are seen in nature. The general mechanism for methyl transfer is a SN2-like nucleophilic attack where the methionine sulfur serves as the leaving group and the methyl group attached to it acts as the electrophile that transfers the methyl group to the enzyme substrate. SAM is converted to S-Adenosyl homocysteine (SAH) during this process. The breaking of the SAM-methyl bond and the formation of the substrate-methyl bond happen nearly simultaneously. These enzymatic reactions are found in many pathways and are implicated in genetic diseases, cancer, and metabolic diseases. Another type of methyl transfer is the radical S-Adenosyl methionine (SAM) which is the methylation of unactivated carbon atoms in primary metabolites, proteins, lipids, and RNA.

<span class="mw-page-title-main">DNA adenine methylase</span> Prokaryotic enzyme

DNA adenine methylase, (Dam) (also site-specific DNA-methyltransferase (adenine-specific), EC 2.1.1.72, modification methylase, restriction-modification system) is an enzyme that adds a methyl group to the adenine of the sequence 5'-GATC-3' in newly synthesized DNA. Immediately after DNA synthesis, the daughter strand remains unmethylated for a short time. It is an orphan methyltransferase that is not part of a restriction-modification system and regulates gene expression. This enzyme catalyses the following chemical reaction

The adaptive response is a form of direct DNA repair in E. coli that protects DNA from damage by external agents or by errors during replication. It is initiated against alkylation, particularly methylation, of guanine or thymine nucleotides or phosphate groups on the sugar-phosphate backbone of DNA. Under sustained exposure to low-level treatment with alkylating mutagens, E. coli can adapt to the presence of the mutagen, rendering subsequent treatment with high doses of the same agent less effective.

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

Fat mass and obesity-associated protein also known as alpha-ketoglutarate-dependent dioxygenase FTO is an enzyme that in humans is encoded by the FTO gene located on chromosome 16. As one homolog in the AlkB family proteins, it is the first mRNA demethylase that has been identified. Certain alleles of the FTO gene appear to be correlated with obesity in humans.

DNA adenine methyltransferase identification, often abbreviated DamID, is a molecular biology protocol used to map the binding sites of DNA- and chromatin-binding proteins in eukaryotes. DamID identifies binding sites by expressing the proposed DNA-binding protein as a fusion protein with DNA methyltransferase. Binding of the protein of interest to DNA localizes the methyltransferase in the region of the binding site. Adenine methylation does not occur naturally in eukaryotes and therefore adenine methylation in any region can be concluded to have been caused by the fusion protein, implying the region is located near a binding site. DamID is an alternate method to ChIP-on-chip or ChIP-seq.

<span class="mw-page-title-main">METTL3</span> Gene encoding part of N6-adenosine-methyltransferase

N6-adenosine-methyltransferase 70 kDa subunit (METTL3) is an enzyme that in humans is encoded by the METTL3 gene.

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

HBx is a hepatitis B viral protein. It is 154 amino acids long and interferes with transcription, signal transduction, cell cycle progress, protein degradation, apoptosis and chromosomal stability in the host. It forms a heterodimeric complex with its cellular target protein, and this interaction dysregulates centrosome dynamics and mitotic spindle formation. It interacts with DDB1 redirecting the ubiquitin ligase activity of the CUL4-DDB1 E3 complexes, which are intimately involved in the intracellular regulation of DNA replication and repair, transcription and signal transduction.

In DNA repair, the Ada regulon is a set of genes whose expression is essential to adaptive response, which is triggered in prokaryotic cells by exposure to sub-lethal doses of alkylating agents. This allows the cells to tolerate the effects of such agents, which are otherwise toxic and mutagenic.

<i>N</i><sup>6</sup>-Methyladenosine Modification in mRNA, DNA

N6-Methyladenosine (m6A) was originally identified and partially characterised in the 1970s, and is an abundant modification in mRNA and DNA. It is found within some viruses, and most eukaryotes including mammals, insects, plants and yeast. It is also found in tRNA, rRNA, and small nuclear RNA (snRNA) as well as several long non-coding RNA, such as Xist.

DNA oxidative demethylase (EC 1.14.11.33, alkylated DNA repair protein, alpha-ketoglutarate-dependent dioxygenase ABH1, alkB (gene)) is an enzyme with systematic name methyl DNA-base, 2-oxoglutarate:oxygen oxidoreductase (formaldehyde-forming). This enzyme catalyses the following chemical reaction

Within the field of molecular biology, the epitranscriptome includes all the biochemical modifications of the RNA within a cell. In analogy to epigenetics that describes "functionally relevant changes to the genome that do not involve a change in the nucleotide sequence", epitranscriptomics involves all functionally relevant changes to the transcriptome that do not involve a change in the ribonucleotide sequence. Thus, the epitranscriptome can be defined as the ensemble of such functionally relevant changes.

<span class="mw-page-title-main">Chuan He</span> Chinese-American chemical biologist

Chuan He is a Chinese-American chemical biologist. He currently serves as the John T. Wilson Distinguished Service Professor at the University of Chicago, and an Investigator of the Howard Hughes Medical Institute. He is best known for his work in discovering and deciphering reversible RNA methylation in post-transcriptional gene expression regulation. He was awarded the 2023 Wolf Prize in Chemistry for his work in discovering and deciphering reversible RNA methylation in post-transcriptional gene expression regulation in addition to his contributions to the invention of TAB-seq, a biochemical method that can map 5-hydroxymethylcytosine (5hmC) at base-resolution genome-wide, as well as hmC-Seal, a method that covalently labels 5hmC for its detection and profiling.

<span class="mw-page-title-main">Epitranscriptomic sequencing</span>

In epitranscriptomic sequencing, most methods focus on either (1) enrichment and purification of the modified RNA molecules before running on the RNA sequencer, or (2) improving or modifying bioinformatics analysis pipelines to call the modification peaks. Most methods have been adapted and optimized for mRNA molecules, except for modified bisulfite sequencing for profiling 5-methylcytidine which was optimized for tRNAs and rRNAs.

<span class="mw-page-title-main">AlkB homolog 5, RNA demethylase</span> Protein-coding gene in the species Homo sapiens

RNA demethylase ALKBH5 is a protein that in humans is encoded by the ALKBH5 gene.

<span class="mw-page-title-main">Alkb homolog 1, histone h2a dioxygenase</span> Protein-coding gene in the species Homo sapiens

AlkB homolog 1, histone H2A dioxygenase is a protein that in humans is encoded by the ALKBH1 gene.

1,<i>N<sup>6</sup></i>-Ethanoadenine Chemical compound

1,N6-Ethanoadenine or epsilonA is a tricyclic derivative of adenine, where an ethylene bridge connect the amine group to the adjacent carbon on the six member ring, to add another five membered ring. This kind of modification can take place as a mutation of DNA. As a DNA modification it is called an etheno (ε) DNA adduct. Chloroacetaldehyde and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) can react with adenine to form 1,N6-ethanoadenine.

References

  1. 1 2 3 4 5 Errol C.Friedberg, Graham c. Walker, Wolfram Siede, Richard D. Wood, Roger A. schultz, Tom Ellenberger, DNA Repair and Mutagenesis, 2nd Edition, ASM press, ISBN   1-55581-319-4
  2. ↑ Yu, Bomina; Hunt, John F. (25 August 2009). "Enzymological and structural studies of the mechanism of promiscuous substrate recognition by the oxidative DNA repair enzyme AlkB". Proceedings of the National Academy of Sciences USA. 106 (34): 14315–14320. Bibcode:2009PNAS..10614315Y. doi: 10.1073/pnas.0812938106 . PMC   2725012 . PMID   19706517.
  3. ↑ Dinglay; et al. (2000). "Defective processing of methylated single-stranded DNA by E. coli alkB mutants". Genes Dev. 14 (16): 2097–2105. doi:10.1101/gad.14.16.2097. PMC   316854 . PMID   10950872.
  4. 1 2 3 Trewick; et al. (2002). "Oxidative demethylation by E. coli alkB directly reverts DNA base damage". Nature. 419 (6903): 174–178. doi:10.1038/nature00908. PMID   12226667. S2CID   4324333.
  5. ↑ Jeanette Ringvoll; et al. (2006). "Repair deficient mice reveal mABH2 as the primary oxidative demethylase for repairing 1meA and 3meC lesions in DNA". The EMBO Journal. 25 (10): 2189–2198. doi:10.1038/sj.emboj.7601109. PMC   1462979 . PMID   16642038.
  6. ↑ Fu, Y; Dai, Q; Zhang, W; Ren, J; Pan, T; He, C (Nov 15, 2010). "The AlkB domain of mammalian ABH8 catalyzes hydroxylation of 5-methoxycarbonylmethyluridine at the wobble position of tRNA". Angewandte Chemie International Edition in English. 49 (47): 8885–8. doi:10.1002/anie.201001242. PMC   3134247 . PMID   20583019.
  7. ↑ van den Born, Erwin; Vågbø, Cathrine B.; Songe-Møller, Lene; Leihne, Vibeke; Lien, Guro F.; Leszczynska, Grazyna; Malkiewicz, Andrzej; Krokan, Hans E.; Kirpekar, Finn; Klungland, Arne; Falnes, Pål Ø. (1 February 2011). "ALKBH8-mediated formation of a novel diastereomeric pair of wobble nucleosides in mammalian tRNA". Nature Communications. 2: 172. Bibcode:2011NatCo...2..172V. doi: 10.1038/ncomms1173 . hdl: 10852/74450 . PMID   21285950.
  8. ↑ Jia, Guifang; Fu, Ye; Zhao, Xu; Dai, Qing; Zheng, Guanqun; Yang, Ying; Yi, Chengqi; Lindahl, Tomas; Pan, Tao; Yang, Yun-Gui; He, Chuan (16 October 2011). "N6-Methyladenosine in nuclear RNA is a major substrate of the obesity-associated FTO". Nature Chemical Biology. 7 (12): 885–887. doi:10.1038/nchembio.687. PMC   3218240 . PMID   22002720.
  9. ↑ Bratlie, Marit S.; Drabløs, Finn (2005-01-03). "Bioinformatic mapping of AlkB homology domains in viruses". BMC Genomics. 6 (1): 1. doi: 10.1186/1471-2164-6-1 . ISSN   1471-2164. PMC   544882 . PMID   15627404.
  10. ↑ Pasin, Fabio; Daròs, José-Antonio; Tzanetakis, Ioannis E. (2022-07-01). "Proteome expansion in the Potyviridae evolutionary radiation". FEMS Microbiology Reviews. 46 (4): fuac011. doi:10.1093/femsre/fuac011. ISSN   1574-6976. PMC   9249622 . PMID   35195244.
  11. 1 2 Yue, Jianying; Wei, Yao; Sun, Zhenqi; Chen, Yahan; Wei, Xuefeng; Wang, Haijuan; Pasin, Fabio; Zhao, Mingmin (October 2022). "AlkB RNA demethylase homologues and N6 -methyladenosine are involved in Potyvirus infection". Molecular Plant Pathology. 23 (10): 1555–1564. doi:10.1111/mpp.13239. ISSN   1364-3703. PMC   9452765 . PMID   35700092.
  12. ↑ Susaimuthu, James; Tzanetakis, Ioannis E.; Gergerich, Rose C.; Martin, Robert R. (February 2008). "A member of a new genus in the Potyviridae infects Rubus". Virus Research. 131 (2): 145–151. doi:10.1016/j.virusres.2007.09.001. PMID   17933412.
  13. ↑ Delaney; et al. (2004). "Mutagenesis, genotoxicity, and repair of 1-methyladenine, 3-alkylcytosines, 1-methylguanine, and 3-methylthymine in alkB Escherichia coli". PNAS. 101 (39): 14051–14056. Bibcode:2004PNAS..10114051D. doi: 10.1073/pnas.0403489101 . PMC   521119 . PMID   15381779.
  14. ↑ Frick; et al. (2007). "Alleviation of 1,N6-ethanoadenine genotoxicity by the Escherichia coli adaptive response protein AlkB". PNAS. 104 (3): 755–760. Bibcode:2007PNAS..104..755F. doi: 10.1073/pnas.0607377104 . PMC   1783386 . PMID   17213319.
  15. ↑ Delaney; et al. (2005). "AlkB reverses etheno DNA lesions caused by lipid oxidation in vitro and in vivo". Nat. Struct. Mol. Biol. 12 (10): 855–860. doi:10.1038/nsmb996. PMID   16200073. S2CID   23235920.
  16. ↑ van den Born, Erwin; Omelchenko, Marina V.; Bekkelund, Anders; Leihne, Vibeke; Koonin, Eugene V.; Dolja, Valerian V.; Falnes, Pål Ø (October 2008). "Viral AlkB proteins repair RNA damage by oxidative demethylation". Nucleic Acids Research. 36 (17): 5451–5461. doi:10.1093/nar/gkn519. ISSN   1362-4962. PMC   2553587 . PMID   18718927.
  17. ↑ Martínez-Pérez, Mireya; Aparicio, Frederic; López-Gresa, Maria Pilar; Bellés, Jose María; Sánchez-Navarro, Jesus A.; Pallás, Vicente (2017-10-03). "Arabidopsis m6A demethylase activity modulates viral infection of a plant virus and the m6A abundance in its genomic RNAs". Proceedings of the National Academy of Sciences of the United States of America. 114 (40): 10755–10760. Bibcode:2017PNAS..11410755M. doi: 10.1073/pnas.1703139114 . ISSN   1091-6490. PMC   5635872 . PMID   28923956.
  18. ↑ Martínez-Pérez, Mireya; Gómez-Mena, Concepción; Alvarado-Marchena, Luis; Nadi, Riad; Micol, José Luis; Pallas, Vicente; Aparicio, Frederic (2021). "The m6A RNA Demethylase ALKBH9B Plays a Critical Role for Vascular Movement of Alfalfa Mosaic Virus in Arabidopsis". Frontiers in Microbiology. 12: 745576. doi: 10.3389/fmicb.2021.745576 . ISSN   1664-302X. PMC   8521051 . PMID   34671333.
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