title: A purified subfragment of yeast Atp11p retains full molecular chaperone activity.
abstract: Atp11p is a molecular chaperone of the mitochondrial matrix that participates in the biogenesis pathway to form F1, the catalytic unit of the ATP synthase. Affinity tag pull-down assays and yeast two-hybrid screens have shown that Atp11p binds to free beta subunits of F1 (Wang, Z. G., and Ackerman, S. H. (2000) J. Biol. Chem. 275, 5767-5772). This binding action prevents the beta subunit from associating with itself in non-productive complexes and fosters the formation of a (alpha beta)3 hexamer. Following the premise that Atp11p action is mediated primarily through a surface (as opposed to specific amino acids, as in an enzyme active site), solving its three-dimensional structure so that we may learn how the shape of the protein influences its function is a high priority. Recombinant yeast Atp11p has proven refractory for such analysis because of the presence of a disordered region in the protein. In this article, we show that removal of 67 residues from the amino terminus of recombinant Atp11p yields a subfragment of the protein (called Atp11pTRNC) that retains molecular chaperone function as determined in vitro with both a surrogate substrate (reduced insulin) and the natural substrate (F1 beta). Moreover, preliminary 15N-1H heteronuclear single quantum coherence spectra obtained with Atp11pTRNC indicate that the truncated protein is well ordered and amenable to structure determination by nuclear magnetic resonance.
authors.affiliation: Department of Surgery, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
authors.forename: Ayana
authors.initials: A
authors.lastname: Hinton
authors.forename: Erik R P
authors.initials: ER
authors.lastname: Zuiderweg
authors.forename: Sharon H
authors.initials: SH
authors.lastname: Ackerman
mesh_terms: D001665:Binding Sites; D002384:Catalysis; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D008957:Models, Genetic; D018832:Molecular Chaperones / Q000737:chemistry* / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000378:metabolism; D013997:Time Factors
chemical_list: C434343:Atp11 protein, S pombe; D018832:Molecular Chaperones; D011994:Recombinant Proteins; D029702:Schizosaccharomyces pombe Proteins; D006180:Proton-Translocating ATPases