title: Genetic analysis of selenocysteine biosynthesis in the archaeon Methanococcus maripaludis.
abstract: In Archaea selenocysteine (Sec) is synthesized in three steps. First seryl-tRNA synthetase acylates tRNA(Sec) with serine to generate Ser-tRNA(Sec). Then phosphoseryl-tRNA(Sec) kinase (PSTK) forms Sep-tRNA(Sec) , which is converted to Sec-tRNA(Sec) by Sep-tRNA:Sec-tRNA synthase (SepSecS) in the presence of selenophosphate produced by selenophosphate synthetase (SelD). A complete in vivo analysis of the archaeal Sec biosynthesis pathway is still unavailable, and the existence of a redundant pathway or of a rescue mechanism based on the conversion of Sep-tRNA(Sec) to Cys-tRNA(Sec) during selenium starvation, cannot be excluded. Here we present a mutational analysis of Sec biosynthesis in Methanococcus maripaludis strain Mm900. Sec formation is abolished upon individually deleting the genes encoding SelD, PSTK or SepSecS; the resulting mutant strains could no longer grow on formate while growth with H(2) + CO(2) remained unaffected. However, deletion of the PSTK and SepSecS genes was not possible unless the selenium-free [NiFe]-hydrogenases Frc and Vhc were expressed. This required the prior deletion of either the gene encoding SelD or that of HrsM, a LysR-type regulator suppressing transcription of the frc and vhc operons in the presence of selenium. These results show that M. maripaludis Mm900 is facultatively selenium-dependent with a single pathway of Sec-tRNA(Sec) formation.
authors.affiliation: Departments of Molecular Biophysics and Biochemistry Chemistry, Yale University, New Haven, CT 06520-8114, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Hohn
authors.forename: Sotiria
authors.initials: S
authors.lastname: Palioura
authors.forename: Dan
authors.initials: D
authors.lastname: Su
authors.forename: Jing
authors.initials: J
authors.lastname: Yuan
authors.forename: Dieter
authors.initials: D
authors.lastname: Söll
mesh_terms: D019843:Archaeal Proteins / Q000235:genetics / Q000378:metabolism; D053898:Biosynthetic Pathways / Q000235:genetics*; D002245:Carbon Dioxide / Q000378:metabolism; D005561:Formates / Q000378:metabolism; D017353:Gene Deletion; D006859:Hydrogen / Q000378:metabolism; D017017:Methanococcus / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D012345:RNA, Transfer, Amino Acid-Specific / Q000378:metabolism*; D017279:Selenocysteine / Q000096:biosynthesis*
chemical_list: D019843:Archaeal Proteins; D005561:Formates; D012345:RNA, Transfer, Amino Acid-Specific; C036928:tRNA, selenocysteine-; D017279:Selenocysteine; C030544:formic acid; D002245:Carbon Dioxide; D006859:Hydrogen