Journal Article

Structural basis for tRNA methylthiolation by the radical SAM enzyme MiaB

Almo, Steven C., Wang, Bo, Yennawar, Neela H., Krebs, Carsten, Esakova, Olga A., Arcinas, Arthur J., Grove, Tyler L., Booker, Squire J.

2021

Overview

Numerous post-transcriptional modifications of transfer RNAs have vital roles in translation. The 2-methylthio- N 6 -isopentenyladenosine (ms 2 i 6 A) modification occurs at position 37 (A37) in transfer RNAs that contain adenine in position 36 of the anticodon, and serves to promote efficient A:U codon–anticodon base-pairing and to prevent unintended base pairing by near cognates, thus enhancing translational fidelity 1 – 4 . The ms 2 i 6 A modification is installed onto isopentenyladenosine (i 6 A) by MiaB, a radical S -adenosylmethionine (SAM) methylthiotransferase. As a radical SAM protein, MiaB contains one [Fe 4 S 4 ] RS cluster used in the reductive cleavage of SAM to form a 5ʹ-deoxyadenosyl 5ʹ-radical, which is responsible for removing the C 2 hydrogen of the substrate 5 . MiaB also contains an auxiliary [Fe 4 S 4 ] aux cluster, which has been implicated 6 – 9 in sulfur transfer to C 2 of i 6 A37. How this transfer takes place is largely unknown. Here we present several structures of MiaB from Bacteroides uniformis . These structures are consistent with a two-step mechanism, in which one molecule of SAM is first used to methylate a bridging µ-sulfido ion of the auxiliary cluster. In the second step, a second SAM molecule is cleaved to a 5ʹ-deoxyadenosyl 5ʹ-radical, which abstracts the C 2 hydrogen of the substrate but only after C 2 has undergone rehybridization from sp 2 to sp 3 . This work advances our understanding of how enzymes functionalize inert C–H bonds with sulfur. Crystal structures reveal the catalytic mechanism through which the radical S -adenosylmethionine enzyme MiaB adds a methylthio group onto tRNA.

• Publisher: Nature Publishing Group UK,Nature Publishing Group
• Subject: 631/45/49/1141 / 631/535/1266 / 82/80 / 82/83 / Adenine / Adenosine / Adenosine - analogs & derivatives / Adenosine - metabolism / Adenosylmethionine / Bacteroides - enzymology / Binding Sites / Biocatalysis / Clusters / Cyclin-dependent kinases / Enzymes / Genetic aspects / Humanities and Social Sciences / Hydrogen / Isopentenyladenosine - metabolism / Methyltransferases - chemistry / Methyltransferases - metabolism / Models, Molecular / multidisciplinary / Oxygen / Physiological aspects / Post-transcription / Protein Domains / Proteins / Radicals / RNA - metabolism / RNA, Transfer - chemistry / RNA, Transfer - metabolism / RNA-Binding Proteins - chemistry / RNA-Binding Proteins - metabolism / S-Adenosylmethionine - metabolism / Science / Science (multidisciplinary) / Structure / Substrate Specificity / Substrates / Sulfhydryl Compounds - metabolism / Sulfur / Sulfurtransferases - chemistry / Sulfurtransferases - metabolism / Transfer RNA / tRNA