Computational design of the sequence and structure of a protein-binding peptide

Abstract

The de novo design of protein-binding peptides is challenging because it requires the identification of both a sequence and a backbone conformation favorable for binding. We used a computational strategy that iterates between structure and sequence optimization to redesign the C-terminal portion of the RGS14 GoLoco motif peptide so that it adopts a new conformation when bound to Gα(i1). An X-ray crystal structure of the redesigned complex closely matches the computational model, with a backbone root-mean-square deviation of 1.1 Å.

Figure 1

Redesigning the RGS14 GoLoco motif. (a) Crystal structure of the wildtype RGS14 GoLoco motif peptide bound to Gα_i1. (b) Model of the redesigned GoLoco motif, GL_helix-4, bound to Gα_i1. Both the wildtype GoLoco motif and GL_helix-4 are shown in cartoon representation with selected side chains displayed and labeled. Gα_i1 is in surface representation with side-chain nitrogens colored blue and side-chain oxygens colored red.

Figure 2

Crystal Structure of GL_helix-4 bound to Ga_i1·GDP. The unbiased 2Fo-Fc electron density (blue mesh, contoured to σ=1.5, see Supplementary Methods), indicates a GoLoco motif peptide C-terminal α-helix (salmon) bound to the all-helical domain of Gα_i1 (gray surface). The well-defined aromatic side chains Phe 525, Phe 531, and Trp 534 establish the correct helical orientation and register.

Figure 3

Model (light and dark green) and crystal structure (salmon and maroon) of GL_helix-4 bound to Gα_i1. The structures were superimposed by minimizing the root mean square deviation between backbone heavy atoms in the all helical domain of Gα_i1. The peptide side chains of Val 524 and Phe 531 pack on either side of Phe 108 from Ga_i1, as predicted. Only the orientation of Trp 534 and the absence of a definable residue 535 in the crystal structure deviate significantly from the predicted model.