A Purified Subfragment of Yeast Atp11p Retains Full Molecular Chaperone Activity

• Published in: The Journal of biological chemistry Vol. 278; no. 36; pp. 34110 - 34113
• Main Authors: Hinton, Ayana, Zuiderweg, Erik R.P., Ackerman, Sharon H.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 05.09.2003; American Society for Biochemistry and Molecular Biology
• Subjects: Binding Sites; Catalysis; Electrophoresis, Polyacrylamide Gel; Hydrogen-Ion Concentration; Magnetic Resonance Spectroscopy; Models, Genetic; Molecular Chaperones - chemistry; Molecular Chaperones - metabolism; Plasmids - metabolism; Protein Binding; Protein Conformation; Protein Structure, Tertiary; Proton-Translocating ATPases - chemistry; Proton-Translocating ATPases - metabolism; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Saccharomyces cerevisiae - enzymology; Schizosaccharomyces pombe Proteins - chemistry; Schizosaccharomyces pombe Proteins - metabolism; Time Factors
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M305353200

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 β subunits of F1 (Wang, Z. G., and Ackerman, S. H. (2000) J. Biol. Chem. 275, 5767–5772). This binding action prevents the β subunit from associating with itself in non-productive complexes and fosters the formation of a (αβ)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 β). 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.