作者: Patrick G. Needham , Hardik J. Patel , Gabriela Chiosis , Patrick H. Thibodeau , Jeffrey L. Brodsky
DOI: 10.1016/J.JMB.2015.04.010
关键词:
摘要: The major cytoplasmic Hsp70 chaperones in the yeast Saccharomyces cerevisiae are Ssa proteins, and much of our understanding biology has emerged from studying ssa mutant strains. For example, Ssa1 catalyzes multiple cellular functions, including protein transport degradation, to this end, ssa1-45 proved invaluable. However, biochemical defects associated with corresponding Ssa1-45 (P417L) unknown. Consequently, we characterized P417L, as well a P417S variant, which corresponds mutation gene encoding mitochondrial Hsp70. We discovered that P417L proteins exhibit accelerated ATPase activity was similar Hsp40-stimulated rate ATP hydrolysis wild-type Ssa1. also found were compromised for peptide binding. These data consistent peptide-stimulated results limited proteolysis experiments, indicated mutants' substrate binding domains highly vulnerable digestion. Defects reactivation heat-denatured luciferase evident. Correspondingly, expressing or only copy temperature sensitive exhibited Ssa1-dependent translocation misfolded degradation. Together, studies suggest structure domain is altered coupling between nucleotide disabled when conserved P417 residue mutated. Our provide new insights into nature many allele.