A Set ofComputationally Designed Orthogonal AntiparallelHomodimers that Expands the Synthetic Coiled-Coil Toolkit

摘要

Molecular engineering of protein assemblies, including the fabrication of nanostructures and synthetic signaling pathways, relies on the availability of modular parts that can be combined to give different structures and functions. Currently, a limited number of well-characterized protein interaction components are available. Coiled-coil interaction modules have been demonstrated to be useful for biomolecular design, and many parallel homodimers and heterodimers are available in the coiled-coil toolkit. In this work, we sought to design a set of orthogonal antiparallel homodimeric coiled coils using a computational approach. There are very few antiparallel homodimers described in the literature, and none have been measured for cross-reactivity. We tested the ability of the distance-dependent statistical potential DFIRE to predict orientation preferences for coiled-coil dimers of known structure. The DFIRE model was then combined with the CLASSY multistate protein design framework to engineer sets of three orthogonal antiparallel homodimeric coiled coils. Experimental measurements confirmed the successful design ofthree peptides that preferentially formed antiparallel homodimersthat, furthermore, did not interact with one additional previouslyreported antiparallel homodimer. Two designed peptides that formedhigher-order structures suggest how future design protocols couldbe improved. The successful designs represent a significant expansionof the existing protein-interaction toolbox for molecular engineers.