Interdomain interactions dictate the function of the Candida albicans Hsp110 protein Msi3

Heat shock proteins of 110 kDa (Hsp110s), a unique class of molecular chaperones, are essential for maintaining protein homeostasis. Hsp110s exhibit a strong chaperone activity preventing protein aggregation (the holdase activity) and also function as the major nucleotide-exchange factor (NEF) for Hsp70 chaperones. Hsp110s contain two functional domains: a nucleotide-binding domain (NBD) and substrate-binding domain (SBD). ATP binding is essential for Hsp110 function and results in close contacts between the NBD and SBD. However, the molecular mechanism of this ATP-induced allosteric coupling remains poorly defined. In this study, we carried out biochemical analysis on Msi3, the sole Hsp110 in Candida albicans, to dissect the unique allosteric coupling of Hsp110s using three mutations affecting the domain-domain interface. All the mutations abolished both the in vivo and in vitro functions of Msi3. While the ATP-bound state was disrupted in all mutants, only mutation of the NBD-SBD/I interfaces showed significant ATPase activity, suggesting that the full-length Hsp110s have an ATPase that is mainly suppressed by NBD-SBD/I contacts. Moreover, the high-affinity ATP binding unexpectedly appears to require these NBD-SBD contacts. Remarkably, the holdase activity was largely intact for all mutants tested while NEF activity was mostly compromised, although both activities strictly depended on the ATP bound state, indicating different requirements for these two activities. Stable peptide substrate binding to Msi3 led to dissociation of the NBD-SBD contacts and compromised interactions with Hsp70. Taken together, our data demonstrate that the exceptionally strong NBD-SBD contacts in Hsp110s dictate the unique allosteric coupling and biochemical activities.

Automated summary

Hsp110s are a unique class of molecular chaperones essential for maintaining protein homeostasis. They have strong chaperone activity preventing protein aggregation and serve as the major nucleotide-exchange factor for Hsp70 chaperones. Hsp110s contain nucleotide-binding and substrate-binding domains, with ATP binding being crucial for their function and leading to close contacts between these domains.