Format

Send to

Choose Destination
Mol Cell. 2011 Apr 22;42(2):250-60. doi: 10.1016/j.molcel.2011.03.010. Epub 2011 Mar 31.

A de novo protein binding pair by computational design and directed evolution.

Author information

1
Department of Biochemistry, University of Washington, Seattle, WA 98195-7350, USA. johnk@ku.edu

Abstract

The de novo design of protein-protein interfaces is a stringent test of our understanding of the principles underlying protein-protein interactions and would enable unique approaches to biological and medical challenges. Here we describe a motif-based method to computationally design protein-protein complexes with native-like interface composition and interaction density. Using this method we designed a pair of proteins, Prb and Pdar, that heterodimerize with a Kd of 130 nM, 1000-fold tighter than any previously designed de novo protein-protein complex. Directed evolution identified two point mutations that improve affinity to 180 pM. Crystal structures of an affinity-matured complex reveal binding is entirely through the designed interface residues. Surprisingly, in the in vitro evolved complex one of the partners is rotated 180° relative to the original design model, yet still maintains the central computationally designed hotspot interaction and preserves the character of many peripheral interactions. This work demonstrates that high-affinity protein interfaces can be created by designing complementary interaction surfaces on two noninteracting partners and underscores remaining challenges.

PMID:
21458342
PMCID:
PMC3102007
DOI:
10.1016/j.molcel.2011.03.010
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center