Cloning and functional expression of a mammalian gene for a peroxisomal sarcosine oxidase

Sarcosine oxidation in mammals occurs via a mitochondrial dehydrogenase closely linked to the electron transport chain, An additional H2O2-producing sarcosine oxidase has now been purified from rabbit kidney, A corresponding cDNA was cloned from rabbit liver and the gene designated sox, This rabbit sox gene encodes a protein of 390 amino acids and a molecular mass of 44 kDa identical to the molecular mass estimated for the purified enzyme, Sequence analysis revealed an N-terminal ADP-beta alpha beta-binding fold, a motif highly conserved in tightly bound flavoproteins, and a C-terminal peroxiso mal targeting signal 1. Sarcosine oxidase from rabbit liver exhibits high sequence homology (25-28% identity) to monomeric bacterial sarcosine oxidases, Both purified sarcosine oxidase and a recombinant fusion protein synthesized in Escherichia coli contain a covalently bound flavin, metabolize sarcosine, L-pipecolic acid, and L-proline, and cross-react with antibodies raised against L-pipecolic acid oxidase from monkey liver, Subcellular fractionation demonstrated that sarcosine oxidase is a peroxisomal enzyme in rabbit kidney, Transfection of human fibroblast cell lines and CV-1 cells (monkey kidney epithelial cells) with the sox cDNA resulted in a peroxisomal localization of sarcosine oxidase and revealed that the import into the peroxisomes is mediated by the peroxisomal targeting signal 1 pathway.