title: Identification and characterization of serine acetyltransferase encoded by the Mycobacterium tuberculosis Rv2335 gene.
abstract: Serine acetyltransferase (CysE) is the first enzyme involved in the two-step enzymatic pathway of L-cysteine biosynthesis in bacteria and plants, but not in humans. CysE catalyzes the biosynthesis of O-acetyl-L-serine and CoA from L-serine (L-Ser) and acetyl-CoA (AcCoA). Mycobacterium tuberculosis (M. tuberculosis) Rv2335 was predicted as the cysE gene encoding serine acetyltransferase. In this study, the M. tuberculosis Rv2335 gene was cloned and the CysE protein was expressed in E. coli BL21 (DE3). The M. tuberculosis CysE protein was purified by Ni(2+) affinity chromatography and confirmed by SDS-PAGE, western blotting and mass spectrometry. The serine acetyltransferase activity of the M. tuberculosis CysE protein was detected using Ellman's reagent. M. tuberculosis CysE displayed optimal activity at pH 7.5 and 37˚C. The Michaelis constant for AcCoA and L-Ser was 0.0513±0.0050 and 0.0264±0.0006 mM, respectively. The maximum velocity (V(max)) for CysE was 0.0073±0.0005 mM/min. The CysE assay and the determination of the kinetic parameters of M. tuberculosis CysE may be helpful for screening its inhibitors in anti-tuberculosis drug discovery.
authors.affiliation: Department of Biotechnology, Dalian Medical University, Dalian 116044, P.R. China.
authors.forename: Juanjuan
authors.initials: J
authors.lastname: Qiu
authors.forename: Daiqing
authors.initials: D
authors.lastname: Wang
authors.forename: Yufang
authors.initials: Y
authors.lastname: Ma
authors.forename: Tao
authors.initials: T
authors.lastname: Jiang
authors.forename: Yi
authors.initials: Y
authors.lastname: Xin
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D003001:Cloning, Molecular; D005798:Genes, Bacterial / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics*; D051050:Serine O-Acetyltransferase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D013696:Temperature; D013997:Time Factors
chemical_list: D001426:Bacterial Proteins; D051050:Serine O-Acetyltransferase