Year: 2011
Type: article
Abstract: Sep-tRNA:Cys-tRNA synthase (SepCysS) catalyzes the sulfhydrylation of tRNA-bound O-phosphoserine (Sep) to form cysteinyl-tRNA(Cys) (Cys-tRNA(Cys)) in methanogens that lack the canonical cysteinyl-tRNA synthetase (CysRS). A crystal structure of the Archaeoglobus fulgidus SepCysS apoenzyme provides information on the binding of the pyridoxal phosphate cofactor as well as on amino acid residues that may be involved in substrate binding. However, the mechanism of sulfur transfer to form cysteine was not known. Using an in vivo Escherichia coli complementation assay, we showed that all three highly conserved Cys residues in SepCysS (Cys(64), Cys(67), and Cys(272) in the Methanocaldococcus jannaschii enzyme) are essential for the sulfhydrylation reaction in vivo. Biochemical and mass spectrometric analysis demonstrated that Cys(64) and Cys(67) form a disulfide linkage and carry a sulfane sulfur in a portion of the enzyme. These results suggest that a persulfide group (containing a sulfane sulfur) is the proximal sulfur donor for cysteine biosynthesis. The presence of Cys(272) increased the amount of sulfane sulfur in SepCysS by 3-fold, suggesting that this Cys residue facilitates the generation of the persulfide group. Based upon these findings, we propose for SepCysS a sulfur relay mechanism that recruits both disulfide and persulfide intermediates.
Source: Journal of Biological Chemistry
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: hybrid
Estimated APC paid: $3,430
Awards: G1239891305, G222768804, G2453936929, G4137159063, G5648016710, SELENOCYSTEINE: BIOSYNTHESIS AND SIGNIFICANCE FOR HYDROGEN UTLIZATION, G6000251361, ESSENTIAL CRYSTEINE METABOLISM IN ARCHAEA, Studies of Transfer RNA, TRANFER RNA, G7725333995, Integrated Technology Resource for Biomedial Glycomics, G8232595177, G9000074214
OpenAlex ID: W2001011202