Open AccessExpression and Purification of the High‐affinity Phosphate Transporter of Saccharomyces cerevisiae
Author(s) -
Berhe Abraham,
Fristedt Ulrika,
Persson Bengt L.
Publication year - 1995
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1995.tb20426.x
Subject(s) - permease , histidine , saccharomyces cerevisiae , affinity chromatography , biochemistry , phosphate , escherichia coli , chimera (genetics) , chemistry , biology , gene , amino acid , enzyme
The plasma membrane high‐affinity phosphate permease of Saccharomyces cerevisiae has been overproduced as a stable membrane‐bound chimeric protein in Escherichia coli. Construction of a chimera between the permease and a peptide containing 10 consecutive histidine residues allowed selective binding of the chimera to a chelating column charged with Ni 2+ , and elution with imidazole in a high state of purity. Approximately 5 mg purified His 10 ‐permease was obtained from 3 g (wet mass) cells. The purified phosphate permease chimera catalyzes uncoupler‐sensitive phosphate transport after reconstitution into proteoliposomes.