A high throughput approach for the generation of orthogonally interacting protein pairs

Authors

Justin Lawrie, University of Nebraska-LincolnFollow
Xi Song, University of Nebraska-Lincoln
Wei Niu, University of Nebraska-LincolnFollow
Jiantao Guo, University of Nebraska-LincolnFollow

Document Type

Article

Date of this Version

1-17-2018

Citation

Scientific Reports 8 (2018), nr 867.

doi 10.1038/s41598-018-19281-6

Comments

© The Author(s) 2018. This article is licensed under a Creative Commons Attribution 4.0 International License,

Abstract

In contrast to the nearly error-free self-assembly of protein architectures in nature, artificial assembly of protein complexes with pre-defined structure and function in vitro is still challenging. To mimic nature’s strategy to construct pre-defined three-dimensional protein architectures, highly specific proteinprotein interacting pairs are needed. Here we report an effort to create an orthogonally interacting protein pair from its parental pair using a bacteria-based in vivo directed evolution strategy. This high throughput approach features a combination of a negative and a positive selection. The newly developed negative selection from this work was used to remove any protein mutants that retain effective interaction with their parents. The positive selection was used to identify mutant pairs that can engage in effective mutual interaction. By using the cohesin-dockerin protein pair that is responsible for the self-assembly of cellulosome as a model system, we demonstrated that a protein pair that is orthogonal to its parent pair could be readily generated using our strategy. This approach could open new avenues to a wide range of protein-based assembly, such as biocatalysis or nanomaterials, with pre-determined architecture and potentially novel functions and properties.