title: A de novo protein binding pair by computational design and directed evolution.
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.
authors: Department of Biochemistry, University of Washington, Seattle, WA 98195-7350, USA. johnk@ku.edu
authors: John
authors: J
authors: Karanicolas
authors: Jacob E
authors: JE
authors: Corn
authors: Irwin
authors: I
authors: Chen
authors: Lukasz A
authors: LA
authors: Joachimiak
authors: Orly
authors: O
authors: Dym
authors: Sun H
authors: SH
authors: Peck
authors: Shira
authors: S
authors: Albeck
authors: Tamar
authors: T
authors: Unger
authors: Wenxin
authors: W
authors: Hu
authors: Gaohua
authors: G
authors: Liu
authors: Scott
authors: S
authors: Delbecq
authors: Gaetano T
authors: GT
authors: Montelione
authors: Clint P
authors: CP
authors: Spiegel
authors: David R
authors: DR
authors: Liu
authors: David
authors: D
authors: Baker
mesh_terms: D001665:Binding Sites; D002623:Chemistry Techniques, Analytical; D017076:Computer-Aided Design; D008958:Models, Molecular; D008970:Molecular Weight; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013499:Surface Properties
chemical_list: D011506:Proteins