Morsomme, Pierre and Chami, Mohamed and Marco, Sergio and Nader, Joseph and Ketchum, Karen A. and Goffeau, André and Rigaud, Jean-Louis. (2002) Characterization of a hyperthermophilic P-type ATPase from Methanococcus jannaschii expressed in yeast. Journal of Biological Chemistry, 277 (33). pp. 29608-29616.
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Abstract
We report on the biochemical and structural properties of a putative P-type H(+)-ATPase, MJ1226p, from the anaerobic hyperthermophilic Archaea Methanococcus jannaschii. An efficient heterologous expression system was developed in Saccharomyces cerevisiae and a four-step purification protocol, using n-dodecyl beta-d-maltoside, led to a homogeneous detergent-solubilized protein fraction with a yield of over 2 mg of protein per liter of culture. The three-dimensional structure of the purified detergent-solubilized protein obtained at 2.4 nm resolution by electron microscopy showed a dimeric organization in which the size and the shape of each monomer was compatible with the reported structures of P-type ATPases. The purified MJ1226p ATPase was inactive at 40 degrees C and was active at elevated temperature reaching high specific activity, up to 180 micromol of P(i) x min(-1) x mg(-1) at 95 degrees C. Maximum ATPase activity was observed at pH 4.2 and required up to 200 mm monovalent salts. The ATPase activity was stable for several days upon storage at 65 degrees C and was highly resistant to urea and guanidine hydrochloride. The protein formed catalytic phosphoenzyme intermediates from MgATP or P(i), a functional characteristic specific of P-type ATPases. The highly purified, homogeneous, stable, and active MJ1226p ATPase provides a new model for further structure-function studies of P-type ATPases.
Faculties and Departments: | 05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Structural Biology (Engel) 05 Faculty of Science > Departement Biozentrum > Services Biozentrum > BioEM Lab (Chami) |
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UniBasel Contributors: | Chami, Mohamed |
Item Type: | Article, refereed |
Article Subtype: | Research Article |
Publisher: | American Society for Biochemistry and Molecular Biology |
ISSN: | 0021-9258 |
e-ISSN: | 1083-351X |
Note: | Publication type according to Uni Basel Research Database: Journal article |
Language: | English |
Identification Number: |
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edoc DOI: | |
Last Modified: | 17 Feb 2023 14:16 |
Deposited On: | 17 Feb 2023 14:16 |
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