A side reaction of alanine racemase: transamination of cycloserine.
Alanine racemase (EC 5.1.1.1) catalyzes the interconversion of alanine enantiomers, and thus represents the first committed step involved in bacterial cell wall biosynthesis. Cycloserine acts as a suicide inhibitor of alanine racemase and as such, serves as an antimicrobial agent. The chemical means by which cycloserine inhibits alanine racemase is unknown. Through spectroscopic assays, we show here evidence of a pyridoxal derivative (arising from either isomer of cycloserine) saturated at the C4' carbon position. We additionally report the L- and D-cycloserine inactivated crystal structures of Bacillus stearothermophilus alanine racemase, which corroborates the spectroscopy via evidence of a 3-hydroxyisoxazole pyridoxamine derivative. Upon the basis of the kinetic and structural properties of both the L- and D-isomers of the inhibitor, we propose a mechanism of alanine racemase inactivation by cycloserine. This pathway involves an initial transamination step followed by tautomerization to form a stable aromatic adduct, a scheme similar to that seen in cycloserine inactivation of aminotransferases.
Department of Biochemistry and Chemistry, Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, Massachusetts 02454, USA.
Timothy D
TD
Fenn
Geoffrey F
GF
Stamper
Anthony A
AA
Morollo
Dagmar
D
Ringe
D000411:Alanine Racemase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D003523:Cycloserine / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D001411:Geobacillus stearothermophilus / Q000201:enzymology; D007700:Kinetics; D008958:Models, Molecular; D011487:Protein Conformation; D013053:Spectrophotometry; D055672:Static Electricity; D013237:Stereoisomerism
D004791:Enzyme Inhibitors; D003523:Cycloserine; D000411:Alanine Racemase