A Yeast Recombinant Aquaporin Mutant That Is Not Expressed or Mistargeted in Xenopus Oocyte Can Be Functionally Analyzed in Reconstituted Proteoliposomes
| Autor: | Stéphane Deschamps, Jean Gouranton, Frédérique Tacnet, Jean-François Hubert, Valérie Lagrée, Françoise Le Cahérec, Isabelle Pellerin, Nathalie Roudier, Daniel Thomas |
|---|---|
| Rok vydání: | 1998 |
| Předmět: |
Proteolipids
Xenopus Molecular Sequence Data Mutant Saccharomyces cerevisiae Aquaporin Biochemistry Ion Channels law.invention law medicine Animals Amino Acid Sequence Cysteine Molecular Biology biology Wild type Mercury Cell Biology Oocyte biology.organism_classification Molecular biology Yeast medicine.anatomical_structure Mutagenesis Recombinant DNA |
| Zdroj: | Journal of Biological Chemistry. 273:12422-12426 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.273.20.12422 |
| Popis: | We have recently identified AQPcic (foraquaporin cicadella), an insect aquaporin found in the digestive tract of homopteran insects and involved in the elimination of water ingested in excess with the dietary sap (Le Caherec, F., Deschamps, S., Delamarche, C., Pellerin, I., Bonnec, G., Guillam, M. T., Gouranton, J., Thomas, D., and Hubert, J. F. (1996) Eur. J. Biochem.241, 707–715). Like many other aquaporins, AQPcic is inhibited by mercury reagents. In this study, we have demonstrated that residue Cys82 is essential for mercury inhibition. Another mutant version of AQPcic (AQP-C134S), expression of which in Xenopus laevis failed to produce an active molecule, was successfully expressed in Saccharomyces cerevisiae. Using stopped-flow analysis of reconstituted proteoliposomes, we demonstrated that the biological activity and Hg sensitivity of yeast-expressed wild type and mutant type AQPcic was readily assessed. Therefore, we propose that the yeast system is a valid alternative to Xenopus oocytes for studying particular mutants of aquaporin. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|