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Protozoan ALKBH8 oxygenases display both DNA repair and tRNA modification activities.

ABSTRACT: The ALKBH family of Fe(II) and 2-oxoglutarate dependent oxygenases comprises enzymes that display sequence homology to AlkB from E. coli, a DNA repair enzyme that uses an oxidative mechanism to dealkylate methyl and etheno adducts on the nucleobases. Humans have nine different ALKBH proteins, ALKBH1-8 and FTO. Mammalian and plant ALKBH8 are tRNA hydroxylases targeting 5-methoxycarbonylmethyl-modified uridine (mcm5U) at the wobble position of tRNAGly(UCC). In contrast, the genomes of some bacteria encode a protein with strong sequence homology to ALKBH8, and robust DNA repair activity was previously demonstrated for one such protein. To further explore this apparent functional duality of the ALKBH8 proteins, we have here enzymatically characterized a panel of such proteins, originating from bacteria, protozoa and mimivirus. All the enzymes showed DNA repair activity in vitro, but, interestingly, two protozoan ALKBH8s also catalyzed wobble uridine modification of tRNA, thus displaying a dual in vitro activity. Also, we found the modification status of tRNAGly(UCC) to be unaltered in an ALKBH8 deficient mutant of Agrobacterium tumefaciens, indicating that bacterial ALKBH8s have a function different from that of their eukaryotic counterparts. The present study provides new insights on the function and evolution of the ALKBH8 family of proteins.

SUBMITTER: Zdzalik D

PROVIDER: S-EPMC4051686 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Protozoan ALKBH8 oxygenases display both DNA repair and tRNA modification activities.

Zdżalik Daria D Vågbø Cathrine B CB Kirpekar Finn F Davydova Erna E Puścian Alicja A Maciejewska Agnieszka M AM Krokan Hans E HE Klungland Arne A Tudek Barbara B van den Born Erwin E Falnes Pål Ø PØ

PloS one 20140610 6

The ALKBH family of Fe(II) and 2-oxoglutarate dependent oxygenases comprises enzymes that display sequence homology to AlkB from E. coli, a DNA repair enzyme that uses an oxidative mechanism to dealkylate methyl and etheno adducts on the nucleobases. Humans have nine different ALKBH proteins, ALKBH1-8 and FTO. Mammalian and plant ALKBH8 are tRNA hydroxylases targeting 5-methoxycarbonylmethyl-modified uridine (mcm5U) at the wobble position of tRNAGly(UCC). In contrast, the genomes of some bacteri ...[more]

PMID: 24914785

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Project description:Tryptophanyl-tRNA synthetase (TrpRS) is an essential enzyme that is recognizably conserved across all forms of life. It is responsible for activating and attaching tryptophan to a cognate tRNA(Trp) molecule for use in protein synthesis. In some eukaryotes this original core function has been supplemented or modified through the addition of extra domains or the expression of variant TrpRS isoforms. The three TrpRS structures from pathogenic protozoa described here represent three illustrations of this malleability in eukaryotes. The Cryptosporidium parvum genome contains a single TrpRS gene, which codes for an N-terminal domain of uncertain function in addition to the conserved core TrpRS domains. Sequence analysis indicates that this extra domain, conserved among several apicomplexans, is related to the editing domain of some AlaRS and ThrRS. The C. parvum enzyme remains fully active in charging tRNA(Trp) after truncation of this extra domain. The crystal structure of the active, truncated enzyme is presented here at 2.4Å resolution. The Trypanosoma brucei genome contains separate cytosolic and mitochondrial isoforms of TrpRS that have diverged in their respective tRNA recognition domains. The crystal structure of the T. brucei cytosolic isoform is presented here at 2.8Å resolution. The Entamoeba histolytica genome contains three sequences that appear to be TrpRS homologs. However one of these, whose structure is presented here at 3.0Å resolution, has lost the active site motifs characteristic of the Class I aminoacyl-tRNA synthetase catalytic domain while retaining the conserved features of a fully formed tRNA(Trp) recognition domain. The biological function of this variant E. histolytica TrpRS remains unknown, but, on the basis of a completely conserved tRNA recognition region and evidence for ATP but not tryptophan binding, it is tempting to speculate that it may perform an editing function. Together with a previously reported structure of an unusual TrpRS from Giardia, these protozoan structures broaden our perspective on the extent of structural variation found in eukaryotic TrpRS homologs.

Dataset Information

Protozoan ALKBH8 oxygenases display both DNA repair and tRNA modification activities.

Publications

Protozoan ALKBH8 oxygenases display both DNA repair and tRNA modification activities.

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