Folding and Misfolding of a PDZ Tandem Repeat

Although the vast majority of the human proteome is represented by multi-domain proteins, the study of multi-domain folding and misfolding is a relatively poorly explored field. The protein Whirlin is a multidomain scaffolding protein expressed in the inner ear. It is characterized by the presence of tandem repeats of PDZ domains. The first two PDZ domains of Whirlin (PDZ1 and PDZ2 - namely P1P2) are structurally close and separated by a disordered short linker. We recently described the folding mechanism of the P1P2 tandem. The difference in thermodynamic stability of the two domains allowed us to selectively unfold one or both PDZ domains and to pinpoint the accumulation of a misfolded intermediate, which we demonstrated to retain physiological binding activity. In this work, we provide an extensive characterization of the folding and unfolding of P1P2. Based on the observed data, we describe an integrated kinetic analysis that satisfactorily fits the experiments and provides a valuable model to interpret multidomain folding. The experimental and analytical approaches described in this study may be of general interest for the interpretation of complex multi-domain protein folding kinetics. (C) 2021 Elsevier Ltd. All rights reserved.

Automated summary

This study elucidates the folding and unfolding mechanisms of the multi-domain protein Whirlin, focusing on the PDZ1 and PDZ2 domains. The difference in thermodynamic stability of these domains leads to the accumulation of a misfolded intermediate. The experimental and analytical approaches presented in this study offer valuable insights for understanding complex multi-domain protein folding kinetics.