Research of Desulfhydrase Involved in L-cysteine Biosynthetic Pathway in Pseudomonas sp / 微生物学通报

The L-cysteine desulfhydrase gene (cd) of Pseudomonas sp.TS1138 was amplified by PCR,and the amplified gene was recombined in the cloning vector pBluescript SKII.The 1.2kb DNA fragment containing cd was sequenced,and its homology with other desulfhydrases was blast; then the cd was cloned into the expression vector pET-21a(+), and afterward expressed by IPTG inducement.The expression protein was purified by Ni-NTA His-Bind Resin.Then the expression protein was identified by the method of activity staining of desulfhydrase, and the characterization of L-cysteine desulfhydrase and the critical role it played in the L-cysteine biosynthetic pathway were discussed.

Purification and Properties of L-Cysteine Synthetase from Pseudomonas / 微生物学通报

The L-ATC hydrolase and L-SCC amidohydrolase which convert L-ATC to L-cysteine in Pseudomonas sp.TS-1138 are purified about 83.9 and 90.3 fold by salting-out method, Sephadex G-75 gel chromatography, DEAE-cellulose 52 ion exchange and Sephadex G-100 gel chromatography, etc. The purified enzyemes are both demonstrated by SDS-PAGE to be a homogeneous protein. Their molecular weight are about 37.5kD and42.8kDa respectively. The optimum reaction temperature are both 35℃, and the optimum pH are 7.0 and 8.0 respectively. The Km of the two enzymes are 0.67 mmol/L and 0.15 mmol/L, and the Vmax are 0.39?10 -3mmol/L?min and 0.42?10 -3mmol/L?min respectively.