Improved RAD51 Binders Through Motif Shuffling Based on the Modularity of BRC Repeats

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 2021 Nov 13

PMID 34772801

Citations 8

Authors

Laurens H Lindenburg

Teodors Pantelejevs

Fabrice Gielen

Pedro Zuazua-Villar

Maren Butz

Eric Rees

Clemens F Kaminski

Jessica A Downs

Marko Hyvonen

Florian Hollfelder

Affiliations

Soon will be listed here.

Abstract

Exchanges of protein sequence modules support leaps in function unavailable through point mutations during evolution. Here we study the role of the two RAD51-interacting modules within the eight binding BRC repeats of BRCA2. We created 64 chimeric repeats by shuffling these modules and measured their binding to RAD51. We found that certain shuffled module combinations were stronger binders than any of the module combinations in the natural repeats. Surprisingly, the contribution from the two modules was poorly correlated with affinities of natural repeats, with a weak BRC8 repeat containing the most effective N-terminal module. The binding of the strongest chimera, BRC8-2, to RAD51 was improved by -2.4 kCal/mol compared to the strongest natural repeat, BRC4. A crystal structure of RAD51:BRC8-2 complex shows an improved interface fit and an extended β-hairpin in this repeat. BRC8-2 was shown to function in human cells, preventing the formation of nuclear RAD51 foci after ionizing radiation.

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