Small molecule inhibition of a Group II chaperonin: Pinpointing a loop region within the equatorial domain as necessary for protein refolding

摘要

The functionality of regions within the equatorial domain of Group II chaperonins is poorly understood. Previously we showed that a 70 amino acid sequence within this domain on the single-subunit recombinant thermosome from Methanocaldococcus jannaschii (rTHS) contains residues directly responsible for refolding protein substrates [L.M. Bergeron, C. Lee, D.S. Clark, Identification of a critical chaperoning region on an archaeal recombinant thermosome, Biochem. Biophys. Res. Commun. 369 (2008) 707-711]. In the present study, 6-aminopenicillanic acid (6-APA) was found to bind to rTHS and inhibit it from refolding proteins. Fluorescence anisotropy was used to measure a 6-APA/rTHS dissociation constant of 17.1 μM and verify that the binding site is within the first 70 amino-terminal rTHS residues. Docking simulations point to a specific loop region at residues 53-57 on rTHS as the most likely binding region. This loop region is located within the oligomeric association sites of the wild-type thermosome. These results implicate a specific equatorial region of Group II chaperonins in the refolding of proteins, and suggest its importance in conformational changes that accompany chaperone function.