title: Crystallization of a mammalian membrane protein overexpressed in Saccharomyces cerevisiae.
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.
authors.affiliation: Unité de Recherche Associée 2096 of the Centre National de la Recherche Scientifique and Service de Biophysique des Fonctions Membranaires, Département de Biologie Joliot Curie, Commissariat à l'Energie Atomique Saclay, 91191 Gif-sur-Yvette Cedex, France.
authors.forename: Marie
authors.initials: M
authors.lastname: Jidenko
authors.forename: Rikke C
authors.initials: RC
authors.lastname: Nielsen
authors.forename: Thomas Lykke-Møller
authors.initials: TL
authors.lastname: Sørensen
authors.forename: Jesper V
authors.initials: JV
authors.lastname: Møller
authors.forename: Marc
authors.initials: M
authors.lastname: le Maire
authors.forename: Poul
authors.initials: P
authors.lastname: Nissen
authors.forename: Christine
authors.initials: C
authors.lastname: Jaxel
mesh_terms: D000818:Animals; D000252:Calcium-Transporting ATPases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002850:Chromatography, Gel; D003460:Crystallization; D018360:Crystallography, X-Ray; D015870:Gene Expression / Q000235:genetics*; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D011817:Rabbits; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases
chemical_list: D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases