title: Compensation for a defective interaction of the hsp70 ssq1 with the mitochondrial Fe-S cluster scaffold isu.
abstract: Ssq1, a specialized yeast mitochondrial Hsp70, plays a critical role in the biogenesis of proteins containing Fe-S clusters through its interaction with Isu, the scaffold on which clusters are built. Two substitutions within the Ssq1 substrate binding cleft, both of which severely reduced affinity for Isu, had very different effects in vivo. Cells expressing Ssq1(F462S), which had no detectable affinity for Isu, are indistinguishable from Deltassq1 cells, underscoring the importance of the Ssq1-Isu1 interaction in vivo. In contrast, cells expressing Ssq1(V472F), whose affinity for Isu is at least 10-fold lower than that of wild-type Ssq1, had only moderately reduced Fe-S enzyme activities and increased iron levels and grew similarly to wild-type cells. Consistent with the reduced affinity for Isu, the ATPase activity of Ssq1(V472F) was stimulated less well than that of Ssq1 upon addition of Isu and Jac1, the J-protein partner of Ssq1. However, higher concentrations of Jac1 or Isu1, which form a stable complex, could compensate for this defect in stimulation of Ssq1(V472F). Expression of Isu1 was up-regulated 10-fold in ssq1(V472F) compared with wild-type cells, suggesting that formation of a Jac1-Isu1 complex can overcome a lowered affinity of Ssq1 for Isu in vivo as well as in vitro.
authors.affiliation: Department of Molecular and Cellular Biology, Faculty of Biotechnology, University of Gdansk, 24 Kladki, 80-822 Gdansk, Poland and Department of Biochemistry, University of Wisconsin, Madison, WI 53706, USA.
authors.forename: Helena
authors.initials: H
authors.lastname: Knieszner
authors.forename: Brenda
authors.initials: B
authors.lastname: Schilke
authors.forename: Rafal
authors.initials: R
authors.lastname: Dutkiewicz
authors.forename: Patrick
authors.initials: P
authors.lastname: D'Silva
authors.forename: Sara
authors.initials: S
authors.lastname: Cheng
authors.forename: Maikke
authors.initials: M
authors.lastname: Ohlson
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Craig
authors.forename: Jaroslaw
authors.initials: J
authors.lastname: Marszalek
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D020816:Amino Acid Motifs; D016880:Anisotropy; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000378:metabolism; D005990:Glycerol / Q000737:chemistry; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D007501:Iron / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000737:chemistry*; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D010641:Phenotype; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D020349:Surface Plasmon Resonance; D013696:Temperature
chemical_list: D005656:Fungal Proteins; D018840:HSP70 Heat-Shock Proteins; C121520:ISU1 protein, S cerevisiae; D007506:Iron-Sulfur Proteins; C115914:JAC1 protein, S cerevisiae; D024101:Mitochondrial Proteins; D018832:Molecular Chaperones; D010455:Peptides; D011994:Recombinant Proteins; C115913:SSQ1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D007501:Iron; D000251:Adenosine Triphosphatases; D005990:Glycerol