Crystallization of a mammalian membrane protein overexpressed in Saccharomyces cerevisiae.
basic_science · Level V
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- Record sourced from PubMed, PMID 16087876.
- Also identified by PMC identifier 1187984.
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Abstract
The Ca2+-ATPase SERCA1a (sarcoplasmic-endoplasmic reticulum Ca2+-ATPase isoform 1a) from rabbit has been overexpressed in Saccharomyces cerevisiae. This membrane protein was purified by avidin agarose affinity chromatography based on natural biotinylation in the expression host, followed by HPLC gel filtration. Both the functional and structural properties of the overexpressed protein validate the method. Thus, calcium-dependent ATPase activity and calcium transport are essentially intact after reconstitution in proteoliposomes. Moreover, the recombinant protein crystallizes in a form that is isomorphous to the native SERCA1a protein from rabbit, and the diffraction properties are similar. This represents a successful crystallization of a mammalian membrane protein derived from a heterologous expression system, and it opens the way for the study of mutant forms of SERCA1a.
Medical subject headings
- Calcium-Transporting ATPases
- Gene Expression
- Saccharomyces cerevisiae