Identification of a Unique Radical S-Adenosylmethionine Methylase Likely Involved in Methanopterin Biosynthesis in Methanocaldococcus jannaschii

dc.contributorVirginia Techen
dc.contributor.authorAllen, Kylie D.en
dc.contributor.authorXu, Huiminen
dc.contributor.authorWhite, Robert H.en
dc.contributor.departmentBiochemistryen
dc.date.accessed2015-10-01en
dc.date.accessioned2015-11-28T21:53:45Zen
dc.date.available2015-11-28T21:53:45Zen
dc.date.issued2014-07-07en
dc.description.abstractMethanopterin (MPT) and its analogs are coenzymes required for methanogenesis and methylotrophy in specialized microorganisms. The methyl groups at C-7 and C-9 of the pterin ring distinguish MPT from all other pterin-containing natural products. However, the enzyme(s) responsible for the addition of these methyl groups has yet to be identified. Here we demonstrate that a putative radical S-adenosyl-L-methionine (SAM) enzyme superfamily member encoded by the MJ0619 gene in the methanogen Methanocaldococcus jannaschii is likely this missing methylase. When MJ0619 was heterologously expressed in Escherichia coli, various methylated pterins were detected, consistent with MJ0619 catalyzing methylation at C-7 and C-9 of 7,8-dihydro-6-hydroxymethylpterin, a common intermediate in both folate and MPT biosynthesis. Site-directed mutagenesis of Cys77 present in the first of two canonical radical SAM CX3CX2C motifs present in MJ0619 did not inhibit C-7 methylation, while mutation of Cys102, found in the other radical SAM amino acid motif, resulted in the loss of C-7 methylation, suggesting that the first motif could be involved in C-9 methylation, while the second motif is required for C-7 methylation. Further experiments demonstrated that the C-7 methyl group is not derived from methionine and that methylation does not require cobalamin. When E. coli cells expressing MJ0619 were grown with deuterium-labeled acetate as the sole carbon source, the resulting methyl group on the pterin was predominantly labeled with three deuteriums. Based on these results, we propose that this archaeal radical SAM methylase employs a previously uncharacterized mechanism for methylation, using methylenetetrahydrofolate as a methyl group donor.en
dc.description.sponsorshipNational Science Foundationen
dc.description.sponsorshipMCB 0722787en
dc.format.mimetypeapplication/pdfen
dc.identifier.citationAllen, Kylie D., et al. (2014). Identification of a Unique Radical S-Adenosylmethionine Methylase Likely Involved in Methanopterin Biosynthesis in Methanocaldococcus jannaschii. Journal of Bacteriology, 196(18), 3315-3323. doi:10.1128/jb.01903-14en
dc.identifier.doihttps://doi.org/10.1128/jb.01903-14en
dc.identifier.issn0021-9193en
dc.identifier.urihttp://hdl.handle.net/10919/64206en
dc.identifier.urlhttp://jb.asm.org/content/196/18/3315en
dc.language.isoenen
dc.publisherAmerican Society for Microbiologyen
dc.rightsIn Copyrighten
dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/en
dc.titleIdentification of a Unique Radical S-Adenosylmethionine Methylase Likely Involved in Methanopterin Biosynthesis in Methanocaldococcus jannaschiien
dc.title.serialJournal of Bacteriologyen
dc.typeArticle - Refereeden
dc.typeDataseten
dc.type.dcmitypeTexten
dc.type.dcmitypeDataseten

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