| First Author | Steegborn C | Year | 1999 |
| Journal | J Biol Chem | Volume | 274 |
| Issue | 18 | Pages | 12675-84 |
| PubMed ID | 10212249 | Mgi Jnum | J:54544 |
| Mgi Id | MGI:1336470 | Doi | 10.1074/jbc.274.18.12675 |
| Citation | Steegborn C, et al. (1999) Kinetics and inhibition of recombinant human cystathionine gamma-lyase. Toward the rational control of transsulfuration. J Biol Chem 274(18):12675-84 |
| abstractText | The gene encoding human cystathionine gamma-lyase was cloned from total cellular Hep G2 RNA. Fusion to a T7 promoter allowed expression in Escherichia coli, representing the first mammalian cystathionine gamma-lyase overproduced in a bacterial system. About 90% of the heterologous gene product was insoluble, and renaturation experiments from purified inclusion bodies met with limited success. About 5 mg/liter culture of human cystathionine gamma-lyase could also be extracted from the soluble lysis fraction, employing a three-step native procedure. While the enzyme showed high gamma-lyase activity toward L-cystathionine (Km = 0.5 mM, Vmax = 2.5 units/mg) with an optimum pH of 8.2, no residual cystathionine beta-lyase behavior and only marginal reactivity toward L-cystine and L-cysteine were detected. Inhibition studies were performed with the mechanism-based inactivators propargylglycine, trifluoroalanine, and aminoethoxyvinylglycine. Propargylglycine inactivated human cystathionine gamma-lyase much more strongly than trifluoroalanine, in agreement with the enzyme's preference for C-gamma-S bonds. Aminoethoxyvinylglycine showed slow and tight binding characteristics with a Ki of 10.5 microM, comparable with its effect on cystathionine beta-lyase. The results have important implications for the design of specific inhibitors for transsulfuration components. |
