title: Expression, purification and characterization of isoforms of peripheral stalk subunits of human V-ATPase.
abstract: The vacuolar-type H+-ATPase (V-ATPase) is a multi-subunit proton pump that is involved in both intra- and extracellular acidification processes throughout human body. Subunits constituting the peripheral stalk of the V-ATPase are known to have several isoforms responsible for tissue/cell specific different physiological roles. To study the different interaction of these isoforms, we expressed and purified the isoforms of human V-ATPase peripheral stalk subunits using Escherichia coli cell-free protein synthesis system: E1, E2, G1, G2, G3, C1, C2, H and N-terminal soluble part of a1 and a2 isoforms. The purification conditions were different depending on the isoforms, maybe reflecting the isoform specific biochemical characteristics. The purified proteins are expected to facilitate further experiments to study about the cell specific interaction and regulation and thus provide insight into physiological meaning of the existence of several isoforms of each subunit in V-ATPase.
authors.affiliation: Department of Biological Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda-shi, Chiba 278-8510, Japan.
authors.forename: Suhaila
authors.initials: S
authors.lastname: Rahman
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Ishizuka-Katsura
authors.forename: Satoshi
authors.initials: S
authors.lastname: Arai
authors.forename: Shinya
authors.initials: S
authors.lastname: Saijo
authors.forename: Ichiro
authors.initials: I
authors.lastname: Yamato
authors.forename: Mitsutoshi
authors.initials: M
authors.lastname: Toyama
authors.forename: Noboru
authors.initials: N
authors.lastname: Ohsawa
authors.forename: Mio
authors.initials: M
authors.lastname: Inoue
authors.forename: Keiko
authors.initials: K
authors.lastname: Honda
authors.forename: Takaho
authors.initials: T
authors.lastname: Terada
authors.forename: Mikako
authors.initials: M
authors.lastname: Shirouzu
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokoyama
authors.forename: So
authors.initials: S
authors.lastname: Iwata
authors.forename: Takeshi
authors.initials: T
authors.lastname: Murata
mesh_terms: D000595:Amino Acid Sequence; D002474:Cell-Free System; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D020033:Protein Isoforms; D055503:Protein Multimerization; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D025262:Vacuolar Proton-Translocating ATPases / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification
chemical_list: D020033:Protein Isoforms; D021122:Protein Subunits; D011994:Recombinant Proteins; D025262:Vacuolar Proton-Translocating ATPases