Chaperone activity of Cyp18 through hydrophobic condensation that enables rescue of transient misfolded molten globule intermediates.
The single-domain cyclophilin 18 (Cyp18) has long been known to function as a peptidyl-prolyl cis/trans isomerase (PPI) and was proposed by us to also function as a chaperone [Freskgard, P.-O., Bergenhem, N., Jonsson, B.-H., Svensson, M., and Carlsson, U. (1992) Science 258, 466-468]. Later several multidomain PPIs were demonstrated to work as both a peptidyl-prolyl cis/trans isomerase and a chaperone. However, the chaperone ability of Cyp18 has been debated. In this work, we add additional results that show that Cyp18 can both accelerate the rate of refolding and increase the yield of native protein during the folding reaction, i.e., function as both a folding catalyst and a chaperone. Refolding experiments were performed using severely destabilized mutants of human carbonic anhydrase II under conditions where the unfolding reaction is significant and a larger fraction of a more destabilized variant populates molten globule-like intermediates during refolding. A correlation of native state protein stability of the substrate protein versus Cyp18 chaperone activity was demonstrated. The induced correction of misfolded conformations by Cyp18 likely functions through rescue from misfolding of transient molten globule intermediates. ANS binding data suggest that the interaction by Cyp18 leads to an early stage condensation of accessible hydrophobic portions of the misfolding-prone protein substrate during folding. The opposite effect was observed for GroEL known as an unfoldase at early stages of refolding. The chaperone effect of Cyp18 was also demonstrated for citrate synthase, suggesting a general chaperone effect of this PPI.
Moparthi
Satish Babu
SB
IFM-Department of Chemistry, Linköping University, Sweden.
Fristedt
Rikard
R
Mishra
Rajesh
R
Almstedt
Karin
K
Karlsson
Martin
M
Hammarström
Per
P
Carlsson
Uno
U
D000817:Anilino Naphthalenesulfonates / Q000737:chemistry / Q000378:metabolism; D024402:Carbonic Anhydrase II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021983:Cyclophilins / Q000737:chemistry* / Q000378:metabolism* / Q000502:physiology; D004795:Enzyme Stability; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D018832:Molecular Chaperones / Q000737:chemistry* / Q000378:metabolism* / Q000502:physiology; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D015854:Up-Regulation
D000817:Anilino Naphthalenesulfonates; D005456:Fluorescent Dyes; D018832:Molecular Chaperones; D011994:Recombinant Proteins; C027132:1-anilino-8-naphthalenesulfonate; D024402:Carbonic Anhydrase II; D021983:Cyclophilins