Sarcosine dehydrogenase (SARDH) is a liver mitochondrial matrix flavoenzyme that catalyzes the oxidative demethylation of sarcosine. SARDH is defective in patients with sarcosinemia. The predicted 918-amino acid SARDH protein contains a putative 22-amino acid mitochondrial targeting sequence, an ADP-binding site, and a stretch of 12 amino acids that matches the covalent flavin-containing peptide from rat liver Sardh. Human SARDH shares 89% amino acid sequence identity with rat liver Sardh and 34% identity with rat liver dimethylglycine dehydrogenase. Northern blot analysis of various human adult and fetal tissues detected a 4-kb SARDH transcript at high levels in adult and fetal liver and at lower levels in adult pancreas and kidney and fetal kidney. The SARDH gene spans at least 75.3 kb and contains 21 exons.