Structural Basis for the BRCA1 Interaction With Branched DNA

Abstract

Abstract : The Breast Cancer Susceptibility gene 1 (BRCA1) encodes an 1863-amino acid protein that plays a central role in the pathogenesis of hereditary breast cancer. The BRCA1 protein contains an N-terminal RING finger, a central region spanning residues 452-1079 that binds to four-way junction DNA, and two C-terminal BRCT domains, which are critical for BRCA1-mediated tumor suppression and are targets for cancer-causing mutations. This project focuses on the elucidation of the structural determinants of the BRCA1-DNA interaction, an essential step towards a deep understanding of the molecular mechanisms of BRCA1 function in DNA repair during cellular physiology and breast cancer pathogenesis. Specifically, the objective of this proposal is to determine the atomic structure of the central BRCA1 protein region in complex with four-way junction DNA, using X-ray crystallography. Numerous attempts to produce large amounts of soluble BRCA1(452-1079) protein in bacteria were unsuccessful. We therefore identified shorter BRCA1 protein fragments that overlap with BRCA1(452-1079), retain the ability to interact with four-way junction DNA, and can be produced in bacterial cells in a soluble form. These protein fragments are suitable for crystallization experiments with four-way junction DNA and these studies are currently in progress.