Molecular basis of cobalamin-dependent RNA modification

TitleMolecular basis of cobalamin-dependent RNA modification
Publication TypeJournal Article
Year of Publication2016
AuthorsDowling D.P, Miles Z.D, Kohrer C., Maiocco S.J, Elliott S.J, Bandarian V., Drennan C.L
JournalNucleic Acids Research
Date Published2016/11
Type of ArticleArticle
ISBN Number0305-1048
Accession NumberWOS:000393817800042
Keywordscrystal-structure; dehalobacter-restrictus; enzymatic-synthesis; epoxyqueuosine reductase; escherichia-coli; guanine transglycosylase; macromolecular crystallography; nucleoside-q; structural basis; tyrosine transfer rna

Queuosine (Q) was discovered in the wobble position of a transfer RNA (tRNA) 47 years ago, yet the final biosynthetic enzyme responsible for Q-maturation, epoxyqueuosine (oQ) reductase (QueG), was only recently identified. QueG is a cobalamin (Cbl)-dependent, [4Fe-4S] cluster-containing protein that produces the hypermodified nucleoside Qin situ on four tRNAs. To understand how QueG is able to perform epoxide reduction, an unprecedented reaction for a Cbl-dependent enzyme, we have determined a series of high resolution structures of QueG from Bacillus subtilis. Our structure of QueG bound to a tRNATyr anticodon stem loop shows how this enzyme uses a HEAT-like domain to recognize the appropriate anticodons and position the hypermodified nucleoside into the enzyme active site. We find Q bound directly above the Cbl, consistent with a reaction mechanism that involves the formation of a covalent Cbl-tRNA intermediate. Using protein film electrochemistry, we show that two [4Fe-4S] clusters adjacent to the Cbl have redox potentials in the range expected for Cbl reduction, suggesting how Cbl can be activated for nucleophilic attack on oQ. Together, these structural and electrochemical data inform our understanding of Cbl dependent nucleic acid modification.

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