Biochemistry Publications
Noncovalent binding of a cyclic peptide inhibitor to the peptidyl-prolyl isomerase Pin1, explored by hydrogen exchange mass spectrometry
Document Type
Article
Publication Date
1-1-2015
Journal
Canadian Journal of Chemistry
Volume
93
Issue
1
First Page
44
Last Page
50
URL with Digital Object Identifier
10.1139/cjc-2014-0230
Abstract
© 2015 Published by NRC Research Press. Pin1 is a peptidyl-prolyl isomerase (PPIase) that plays a central role in eukaryotic cell cycle regulation, making this protein an interesting target for cancer therapy. Pin1 exhibits high specificity for substrates where proline is preceded by phosphoserine or phosphothreonine. The protein comprises an N-terminal WW (tryptophan-tryptophan) domain and a C-terminal PPIase domain. The cyclic peptide [CRYPEVEIC] (square brackets are used to denote the cyclic structure) represents a lead compound for a new class of nonphosphorylated Pin1 inhibitors. Unfortunately, it has not been possible thus far to characterize the Pin1-[CRYPEVEIC] complex by X-ray crystallography. Thus, the exact binding mode remains unknown. The current work employs hydrogen/deuterium exchange mass spectrometry for gaining insights into the Pin1-[CRYPEVEIC] interactions. The WW domain shows extensive conformational dynamics, both in the presence and in the absence of ligand. In contrast, profound changes in deuteration kinetics are observed in the PPIase domain after the addition of [CRYPEVEIC]. The secondary structure elements 2,3, and 4 exhibit markedly reduced deuteration, consistent with their postulated involvement in ligand binding. Unexpectedly, [CRYPEVEIC] destabilizes the range of residues 61-86, a segment that comprises basic side chains that normally interact with the substrate phosphate. This destabilization is likely caused by steric clashes with Y3 or E5 of the inhibitor. Ligand-induced destabilization has previously been reported for a few other proteins, but effects of this type are not very common. Our findings suggest that future crystallization trials on Pin1 variants deleted for residues in the 61-86 range might provide a path towards high-resolution X-ray structures of Pin1 bound to cyclic peptide inhibitors.