Deciphering the protein-protein specificity code

A fundamental challenge in the field of signal transduction is to decipher how protein structure encodes interaction specificity among members of large families. This paper presents a multidisciplinary approach that combines computations with experiments to understand and redesign the selective recognition of G-proteins by the RGS protein family.

HFSP Cross-Disciplinary Fellow Mickey Kosloff and colleagues
authored on Wed, 27 July 2011

Protein-protein interactions between members of large families can be tailored to their function with either broad or narrow specificity. Understanding the structural and molecular basis for such specificity is a major challenge in biology, as well as in drug design. Yet, beyond single representative examples, little is known of how specificity is determined among large protein families and, in particular, those involved in signal transduction.

This paper describes an approach to address this challenge. As a model system to study specificity, the authors investigated the interactions of heterotrimeric G-proteins with Regulators of G-protein Signaling (RGS proteins). RGS proteins are responsible for turning G-proteins "off". In multiple cascades across most human tissues, RGS proteins determine the duration of the G-protein coupled signaling. RGS proteins have also been implicated in a wide range of human pathologies and are promising drug targets, both as primary targets and as complements to drugs that target other components in G-protein signaling (such as G-protein Coupled Receptors).

To understand how the structure of RGS proteins encodes both their common ability to inactivate G-proteins, and also their selective G-protein recognition, Mickey Kosloff and colleagues integrated structure-based energy calculations with biochemical measurements of RGS protein activity. Using a consensus approach across the eight available RGS-domain/G-protein crystal structures, they established a structure-to-sequence map predicting which RGS residues are essential for function and which RGS residues can modulate specific interactions with the cognate G-protein. This map revealed that, in addition to previously identified conserved residues, RGS proteins contain another group of variable "Modulatory Residues", which reside at the periphery of the RGS-domain/G-protein interface and fine-tune G-protein recognition. Mutations of Modulatory Residues in high-activity RGS proteins impaired RGS function, whereas redesign of low-activity RGS proteins in critical Modulatory positions yielded complete gain-of-function RGS mutants. Therefore, RGS proteins combine a conserved core interface with peripheral Modulatory Residues to selectively optimize G-protein recognition and inactivation.

Finally, the authors applied this approach to a completely different system - the interactions of colicin E7 with its inhibitory immunity proteins, a well-established model for studying protein-protein interaction specificity. Thereby they showed the generality of this structure-based method and its potential use in a scalable "bottom-up" approach to study the structural basis for the "wiring" of signal transduction networks.

Figure Legend: (A) A section of an energy-based map of RGS proteins that identified residues that are either “Significant and Conserved” and underlie common RGS function (marked with red stars above the alignment) or previously unidentified “Modulatory Positions” that are variable yet determine RGS specificity towards G-proteins (marked with purple triangles). (B) Redesign of high-activity RGS proteins in Modulatory Positions resulted in complete loss-of-function. (C) Redesign of low-activity RGS proteins in critical Modulatory Positions achieved full gain-of-function.


Integrating energy calculations with functional assays to decipher the specificity of G protein-RGS protein interactions. Mickey Kosloff, Amanda M. Travis, Dustin E. Bosch, David P. Siderovski and Vadim Y. Arshavsky. Nat. Struct. Mol. Biol. (2011) 18 (7): 846-853.

Pubmed link