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Steegborn, Clemens ; Clausen, Tim ; Sondermann, Peter ; Jacob, Uwe ; Worbs, Michael ; Marinkovic, Snezan ; Huber, Robert ; Wahl, Markus C.:
Kinetics and inhibition of recombinant human cystathionine gamma-lyase : Toward the rational control of transsulfuration.
In: The Journal of Biological Chemistry.
Bd. 274
(1999)
Heft 18
.
- S. 12675-12684.
ISSN 1083-351X
DOI: https://doi.org/10.1074/jbc.274.18.12675
Abstract
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.
Weitere Angaben
| Publikationsform: | Artikel in einer Zeitschrift |
|---|---|
| Begutachteter Beitrag: | Ja |
| Zusätzliche Informationen: | PubMed-ID: 10212249 |
| Institutionen der Universität: | Fakultäten > Fakultät für Biologie, Chemie und Geowissenschaften > Fachgruppe Chemie > Lehrstuhl Biochemie > Lehrstuhl Biochemie I - Proteinbiochemie der Signaltransduktion - Univ.-Prof. Dr. Clemens Steegborn Fakultäten Fakultäten > Fakultät für Biologie, Chemie und Geowissenschaften Fakultäten > Fakultät für Biologie, Chemie und Geowissenschaften > Fachgruppe Chemie Fakultäten > Fakultät für Biologie, Chemie und Geowissenschaften > Fachgruppe Chemie > Lehrstuhl Biochemie |
| Titel an der UBT entstanden: | Nein |
| Themengebiete aus DDC: | 500 Naturwissenschaften und Mathematik > 540 Chemie |
| Eingestellt am: | 13 Apr 2015 06:22 |
| Letzte Änderung: | 16 Jun 2023 07:43 |
| URI: | https://eref.uni-bayreuth.de/id/eprint/10001 |