Monomeric sarcosine oxidase: structure of a covalently flavinylated amine oxidizing enzyme

Background: Monomeric sarcosine oxidases (MSOXs) are among the simplest mem bers of a recently recognized family of eukaryotic and prokaryotic enzymes that catalyze similar oxidative reactions with various secondary or tertiar y amino acids and contain covalently bound flavins. Other members of this f amily include heterotetrameric sarcosine oxidase, N-methyltryptophan oxidas e and pipecolate oxidase. Mammalian sarcosine dehydrogenase and dimethylgly cine dehydrogenase may be more distantly related family members, Results: The X-ray crystal structure of MSOX from Bacillus sp. B-0618, expr essed in Escherichia coli, has been solved at 2.0 Angstrom resolution by mu ltiwavelength anomalous dispersion (MAD) from crystals of the selenomethion ine-substituted enzyme. Fourteen selenium sites, belonging to two MSOX mole cules in the asymmetric unit, were used for MAD phasing and to define the l ocal twofold symmetry axis for electron-density averaging. The structures o f the native enzyme and of two enzyme-inhibitor complexes were also determi ned. Conclusions: MSOX is a two-domain protein with an overall topology most sim ilar to that of D-amino acid oxidase, with which it shares 14% sequence ide ntity. The flavin ring is located in a very basic environment, making conta ct with sidechains of arginine, lysine, histidine and the N-terminal end of a helix dipole, The flavin is covalently attached through an 8 alpha-S-cys teinyl linkage to Cys315 of the catalytic domain. Covalent attachment is pr obably self-catalyzed through interactions with the positive sidechains and the helix dipole. Substrate binding is probably stabilized by hydrogen bon ds between the substrate carboxylate and two basic sidechains, Arg52 and Ly s348, located above the re face of the flavin ring.