SIGNIFICANCE OF CHAPERONIN-10-MEDIATED INHIBITION OF ATP HYDROLYSIS BY CHAPERONIN-60

Chaperonins are essential for the folding of proteins in bacteria, mit ochondria, and chloroplasts, We have functionally characterized the ye ast mitochondrial chaperonins hsp60 and hsp10, In the presence of ADP, one molecule of hsp10 binds to hsp60 with an apparent K-d of 0.9 nM a nd a second molecule of hsp10 binds with a K-d of 24 nM, In the presen ce of ATP, the purified yeast chaperonins mediate the refolding of mit ochondrial malate dehydrogenase. Hsp10 inhibits the ATPase activity of hsp60 by about 40%, Hsp10(P36H) is a point mutant of hsp10 that confe rs temperature-sensitive growth to yeast, Consistent with the in vivo phenotype, refolding of mitochondrial malate dehydrogenase In the pres ence of purified hsp10(P36H) and hsp60 is reduced at 25 degrees C and abolished at 30 degrees C, The affinity of hsp10(P36H) to hsp60 as wel l as to Escherichia coli GroEL Is reduced, However, this decrease in a ffinity does not correlate with the functional defect, because hsp10(P 36H) fully assists the GroEL-mediated refolding of malate dehydrogenas e at 30 degrees C, Refolding activity, rather, correlates with the abi lity of hsp10(P36H) to Inhibit the ATPase of GroEL but not that of hsp 60, Based on our findings, we propose that the inhibition of ATP hydro lysis is mechanistically coupled to chaperonin-mediated protein foldin g.