Identification of a stable flavin-thiolate adduct in heterotetrameric sarcosine oxidase.
Heterotetrameric sarcosine oxidase (TSOX) is a complex bifunctional flavoenzyme that contains two flavins. Most of the FMN in recombinant TSOX is present as a covalent adduct with an endogenous ligand. Enzyme denaturation disrupts the adduct, accompanied by release of a stoichiometric amount of sulfide. Enzyme containing>or=90% unmodified FMN is prepared by displacement of the endogenous ligand with sulfite, a less tightly bound competing ligand. Reaction of adduct-depleted TSOX with sodium sulfide produces a stable complex that resembles the endogenous TSOX adduct and known 4a-S-cysteinyl flavin adducts. The results provide definitive evidence for sulfide as the endogenous TSOX ligand and strongly suggest that the modified FMN is a 4a-sulfide adduct. A comparable reaction with sodium sulfide is not detected with other flavoprotein oxidases. A model of the postulated TSOX adduct suggests that it is stabilized by nearby residues that may be important in the electron transferase/oxidase function of the coenzyme.
Hynson
Robert M G
RM
Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA 19102, USA.
Mathews
F Scott
FS
Schuman Jorns
Marilyn
M
D004228:Dithionitrobenzoic Acid / Q000378:metabolism; D005486:Flavin Mononucleotide / Q000737:chemistry* / Q000378:metabolism*; D008741:Methyl Methanesulfonate / Q000031:analogs & derivatives / Q000378:metabolism; D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D011549:Pseudomonas / Q000201:enzymology*; D050879:Sarcosine Oxidase / Q000737:chemistry* / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000737:chemistry*; D013440:Sulfides / Q000378:metabolism; D013447:Sulfites / Q000378:metabolism
D013438:Sulfhydryl Compounds; D013440:Sulfides; D013447:Sulfites; C014674:methyl methanethiosulfonate; D005486:Flavin Mononucleotide; D004228:Dithionitrobenzoic Acid; D008741:Methyl Methanesulfonate; D050879:Sarcosine Oxidase; C033479:sodium sulfide