Transient Domain Interactions Enhance the Affinity of the Mitotic Regulator Pin1 toward Phosphorylated Peptide Ligands

Abstract

The mitotic regulator Pin1 plays an important role in protein quality control and age-related medical conditions such as Alzheimer disease and Parkinson disease. Although its cellular role has been thoroughly investigated during the past decade, the molecular mechanisms underlying its function remain elusive. We provide evidence for interactions between the two domains of Pin1. Several residues displayed unequivocal peak splits in nuclear magnetic resonance spectra, indicative of two different conformational states in equilibrium. Pareto analysis of paramagnetic relaxation enhancement data demonstrates that the two domains approach each other upon addition of a nonpeptidic ligand. Titration experiments with phosphorylated peptides monitored by fluorescence anisotropy and chemical shift perturbation indicate that domain interactions increase Pin1's affinity toward peptide ligands. We propose this interplay of the domains and ligands to be a general mechanism for a large class of two-domain proteins.

Investigating dynamic interdomain allostery in Pin1

“…7. Yet, other substrate sites may increase interdomain contact, as suggested by FFpSPR and the PEG400 studies of Bayer and co-workers (Matena et al 2013). To be more definitive, we need a more quantitative analysis (e.g.…”

“…A basic MWC premise is a pre-existing equilibrium between distinct apo protein conformations. Evidence for this equilibrium in Pin1 comes from: (i) its solution structure determination, which first suggested conformational selection in Pin1 (Bayer et al 2003); (ii) our findings that loop I of the apo Pin1-WW domain already samples the conformations of the pCdc25C-bound form (Morcos et al 2010; Peng et al 2007); (iii) and the recent studies of Pin1 binding to PEG400, a small non-substrate molecule that consistently occupies the interdomain space in the Pin1 x-ray structures (Matena et al 2013). …”

Structural biology of transcriptional regulation in the c-Myc network

“…The dynamics observed in the NMR spectroscopy structure indicate that the flexible linker and the conformational flexibility are of importance for the function of Pin1 catalysis, and recent studies show that domain interactions increase the affinity of Pin1 for peptide ligands (Labeikovsky, Eisenmesser et al 2007;Matena, Sinnen et al 2013). Pin1 is overexpressed in 38 of 60 tumor types examined (Ayala, Wang et al 2003;Bao, Kimzey et al 2004) and Cyclin D1 is overexpressed in many breast cancer tumors (Bartkova, Lukas et al 1994).…”

Structure and function of the human parvulins Pin1 and Par14/17

“…Furthermore, a negative allosteric regulation of the activity of Pin1 upon inter-domain contact has been reported . The affinity of Pin1 towards substrate ligands, which is mainly attributed to the binding affinity of the WW-domain, is enhanced during inter-domain contact (Matena et al, 2013). However, these in-vitro data still have to be validated for the in vivo behavior of hPin1.…”