Biochemical and genetic studies define the functions of methylthiotransferases in methanogenic and methanotrophic archaea
Abstract
Methylthiotransferases (MTTases) are radical S-adenosylmethionine (SAM) enzymes that catalyze the addition of a methylthio (-SCH3) group to an unreactive carbon center. These enzymes are responsible for the production of 2-methylthioadenosine (ms2A) derivatives found at position A37 of select tRNAs in all domains of life. Additionally, some bacteria contain the RimO MTTase that catalyzes the methylthiolation of the S12 ribosomal protein. Although the functions of MTTases in bacteria and eukaryotes have been established via detailed genetic and biochemical studies, MTTases from the archaeal domain of life are understudied and the substrate specificity determinants of MTTases remain unclear. Here, we report the in vitro enzymatic activities of an MTTase (C4B56_06395) from a thermophilic Ca. Methanophagales anaerobic methanotroph (ANME) as well as the MTTase from a hyperthermophilic methanogen – MJ0867 from Methanocaldococcus jannaschii. Both enzymes catalyze the methylthiolation of N6-threonylcarbamoyladenosine (t6A) and N6-hydroxynorvalylcarbamoyladenosine (hn6A) residues to produce 2-methylthio-N6-threonylcarbamoyladenosine (ms2t6A) and 2-methylthio-N6-hydroxynorvalylcarbamoyladenosine (ms2hn6A), respectively. To further assess the function of archaeal MTTases, we analyzed select tRNA modifications in a model methanogen – Methanosarcina acetivorans – and generated a deletion of the MTTase-encoding gene (MA1153). We found that M. acetivorans produces ms2hn6A in exponential phase of growth, but does not produce ms2t6A in detectable amounts.more »
- Authors:
-
- Virginia Polytechnic Institute and State University (Virginia Tech), Blacksburg, VA (United States); University of Florida, Gainesville, FL (United States)
- Virginia Polytechnic Institute and State University (Virginia Tech), Blacksburg, VA (United States)
- Publication Date:
- Research Org.:
- Virginia Polytechnic Institute and State University (Virginia Tech), Blacksburg, VA (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Basic Energy Sciences (BES). Chemical Sciences, Geosciences & Biosciences Division (CSGB)
- OSTI Identifier:
- 2228923
- Grant/Contract Number:
- SC0022338
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Frontiers in Microbiology
- Additional Journal Information:
- Journal Volume: 14; Journal ID: ISSN 1664-302X
- Publisher:
- Frontiers Research Foundation
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; methanogens; ANME archaea; methylthiotransferases; radical SAM enzymes; tRNA modifications
Citation Formats
Boswinkle, Kaleb, Dinh, Thuc-Anh, and Allen, Kylie D. Biochemical and genetic studies define the functions of methylthiotransferases in methanogenic and methanotrophic archaea. United States: N. p., 2023.
Web. doi:10.3389/fmicb.2023.1304671.
Boswinkle, Kaleb, Dinh, Thuc-Anh, & Allen, Kylie D. Biochemical and genetic studies define the functions of methylthiotransferases in methanogenic and methanotrophic archaea. United States. https://doi.org/10.3389/fmicb.2023.1304671
Boswinkle, Kaleb, Dinh, Thuc-Anh, and Allen, Kylie D. Thu .
"Biochemical and genetic studies define the functions of methylthiotransferases in methanogenic and methanotrophic archaea". United States. https://doi.org/10.3389/fmicb.2023.1304671. https://www.osti.gov/servlets/purl/2228923.
@article{osti_2228923,
title = {Biochemical and genetic studies define the functions of methylthiotransferases in methanogenic and methanotrophic archaea},
author = {Boswinkle, Kaleb and Dinh, Thuc-Anh and Allen, Kylie D.},
abstractNote = {Methylthiotransferases (MTTases) are radical S-adenosylmethionine (SAM) enzymes that catalyze the addition of a methylthio (-SCH3) group to an unreactive carbon center. These enzymes are responsible for the production of 2-methylthioadenosine (ms2A) derivatives found at position A37 of select tRNAs in all domains of life. Additionally, some bacteria contain the RimO MTTase that catalyzes the methylthiolation of the S12 ribosomal protein. Although the functions of MTTases in bacteria and eukaryotes have been established via detailed genetic and biochemical studies, MTTases from the archaeal domain of life are understudied and the substrate specificity determinants of MTTases remain unclear. Here, we report the in vitro enzymatic activities of an MTTase (C4B56_06395) from a thermophilic Ca. Methanophagales anaerobic methanotroph (ANME) as well as the MTTase from a hyperthermophilic methanogen – MJ0867 from Methanocaldococcus jannaschii. Both enzymes catalyze the methylthiolation of N6-threonylcarbamoyladenosine (t6A) and N6-hydroxynorvalylcarbamoyladenosine (hn6A) residues to produce 2-methylthio-N6-threonylcarbamoyladenosine (ms2t6A) and 2-methylthio-N6-hydroxynorvalylcarbamoyladenosine (ms2hn6A), respectively. To further assess the function of archaeal MTTases, we analyzed select tRNA modifications in a model methanogen – Methanosarcina acetivorans – and generated a deletion of the MTTase-encoding gene (MA1153). We found that M. acetivorans produces ms2hn6A in exponential phase of growth, but does not produce ms2t6A in detectable amounts. Upon deletion of MA1153, the ms2A modification was absent, thus confirming the function of MtaB-family MTTases in generating ms2hn6A modified nucleosides in select tRNAs.},
doi = {10.3389/fmicb.2023.1304671},
journal = {Frontiers in Microbiology},
number = ,
volume = 14,
place = {United States},
year = {Thu Nov 23 00:00:00 EST 2023},
month = {Thu Nov 23 00:00:00 EST 2023}
}
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