Rational design using sequence information only produces a peptide that binds to the intrinsically disordered region of p53

Kiyoto Kamagata,Yuji Itoh,Tomoshi Kameda,Agato Murata,Eriko Mano,Ryo Kitahara,Takuro Wakamoto,Hiroto Takahashi,Saori Kanbayashi

doi:10.1038/s41598-019-44688-0

Abstract

Intrinsically disordered regions (IDRs) of proteins are involved in many diseases. The rational drug design against disease-mediating proteins is often based on the 3D structure; however, the flexible structure of IDRs hinders the use of such structure-based design methods. Here, we developed a rational design method to obtain a peptide that can bind an IDR using only sequence information based on the statistical contact energy of amino acid pairs. We applied the method to the disordered C-terminal domain of the tumor suppressor p53. Titration experiments revealed that one of the designed peptides, DP6, has a druggable affinity of ~1 μM to the p53 C-terminal domain. NMR spectroscopy and molecular dynamics simulation revealed that DP6 selectively binds to the vicinity of the target sequence in the C-terminal domain of p53. DP6 inhibits the nonspecific DNA binding of a tetrameric form of the p53 C-terminal domain, but does not significantly affect the specific DNA binding of a tetrameric form of the p53 core domain. Single-molecule measurements revealed that DP6 retards the 1D sliding of p53 along DNA, implying modulation of the target searching of p53. Statistical potential-based design may be useful in designing peptides that target IDRs for therapeutic purposes.

Highlights

Disordered regions (IDRs) of proteins lack a defined 3D structure
The apparent dissociation constant of the designed peptides (KD) with the CT peptide ranged from 1.2 ± 0.8 μM to 550 ± 10 μM in the absence of salt, and DP6 showed the strongest binding to the CT domain (Fig. 1b and Table 1)
We developed the design method of peptides that can bind Intrinsically disordered regions (IDRs) using only the IDR’s sequence information without its structural information

Summary

Introduction

Disordered regions (IDRs) of proteins lack a defined 3D structure. Approximately 10–35% of prokaryotic and 15–45% of eukaryotic proteins are estimated to contain IDRs1. Gabizon et al identified several peptides that bind to the tetrameric form of the disordered CT domain of p53 by screening peptides derived from natural proteins bound to p5329. A series of experiments including NMR, molecular dynamics (MD) simulations, ensemble titration, and single-molecule fluorescence demonstrated that one designed peptide with the highest affinity can modulate the nonspecific DNA binding of p53 and the 1D sliding of p53 along DNA.

Objectives

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Results

Conclusion