期刊
BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES
卷 1818, 期 5, 页码 1342-1350出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbamem.2012.02.004
关键词
Nitrite transport; Dequenching assay; Acridine orange; Solid supported membrane; Transport mechanism
资金
- Max Planck Society
- Deutsche Forschungsgemeinschaft [SFB 807]
- European Union (E-MeP)
In anaerobically grown bacteria, transport of nitrite is catalyzed by an integral membrane protein of the form ate-nitrite transporter family, NirC, which in Salmonella typhimurium plays a critical role in intracellular virulence. We present a functional characterization of the S. typhimurium nitrite transporter StmNirC in native membrane vesicles as well as purified and reconstituted into proteoliposomes. Using an electrophysiological technique based on solid supported membranes, we show nitrite induced translocation of negative charges into proteoliposomes reconstituted with purified StmNirC. These data demonstrate the electrogenicity of StmNirC and its substrate specificity for nitrite. Monitoring changes in Delta pH on everted membrane vesicles containing overexpressed StmNirC using acridine orange as a pH indicator we demonstrate that StmNirC acts as a secondary active transporter. It promotes low affinity transport of nitrite coupled to H+ antiport with a pH independent profile in the pH range from 6 to 8. In addition to nitrite also nitrate is transported by StmNirC, but with reduced flux and complete absence of proton antiport activity. Taken together, these data suggest a bispecific anion selectivity of StmNirC with an ion specific transport mode. This may play a role in regulating nitrite transport under physiological conditions. (C) 2012 Elsevier B.V. All rights reserved.
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