Programmable design of orthogonal protein heterodimers

Zibo Chen, Scott E. Boyken, Mengxuan Jia, Florian Busch, David Flores-Solis, Matthew J. Bick, Peilong Lu, Zachary L. VanAernum, Aniruddha Sahasrabuddhe, Robert A. Langan, Sherry Bermeo, T. J. Brunette, Vikram Khipple Mulligan, Lauren P. Carter, Frank DiMaio, Nikolaos G. Sgourakis, Vicki H. Wysocki and David Baker ()
Additional contact information
Zibo Chen: University of Washington
Scott E. Boyken: University of Washington
Mengxuan Jia: The Ohio State University
Florian Busch: The Ohio State University
David Flores-Solis: University of California Santa Cruz
Matthew J. Bick: University of Washington
Peilong Lu: University of Washington
Zachary L. VanAernum: The Ohio State University
Aniruddha Sahasrabuddhe: The Ohio State University
Robert A. Langan: University of Washington
Sherry Bermeo: University of Washington
T. J. Brunette: University of Washington
Vikram Khipple Mulligan: University of Washington
Lauren P. Carter: University of Washington
Frank DiMaio: University of Washington
Nikolaos G. Sgourakis: University of California Santa Cruz
Vicki H. Wysocki: The Ohio State University
David Baker: University of Washington

Nature, 2019, vol. 565, issue 7737, 106-111

Abstract: Abstract Specificity of interactions between two DNA strands, or between protein and DNA, is often achieved by varying bases or side chains coming off the DNA or protein backbone—for example, the bases participating in Watson–Crick pairing in the double helix, or the side chains contacting DNA in TALEN–DNA complexes. By contrast, specificity of protein–protein interactions usually involves backbone shape complementarity1, which is less modular and hence harder to generalize. Coiled-coil heterodimers are an exception, but the restricted geometry of interactions across the heterodimer interface (primarily at the heptad a and d positions2) limits the number of orthogonal pairs that can be created simply by varying side-chain interactions3,4. Here we show that protein–protein interaction specificity can be achieved using extensive and modular side-chain hydrogen-bond networks. We used the Crick generating equations5 to produce millions of four-helix backbones with varying degrees of supercoiling around a central axis, identified those accommodating extensive hydrogen-bond networks, and used Rosetta to connect pairs of helices with short loops and to optimize the remainder of the sequence. Of 97 such designs expressed in Escherichia coli, 65 formed constitutive heterodimers, and the crystal structures of four designs were in close agreement with the computational models and confirmed the designed hydrogen-bond networks. In cells, six heterodimers were fully orthogonal, and in vitro—following mixing of 32 chains from 16 heterodimer designs, denaturation in 5 M guanidine hydrochloride and reannealing—almost all of the interactions observed by native mass spectrometry were between the designed cognate pairs. The ability to design orthogonal protein heterodimers should enable sophisticated protein-based control logic for synthetic biology, and illustrates that nature has not fully explored the possibilities for programmable biomolecular interaction modalities.

Date: 2019
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DOI: 10.1038/s41586-018-0802-y

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