Structural basis for the BRCA1 BRCT interaction with the proteins ATRIP and BAAT1

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• The breast and ovarian cancer susceptibility protein 1 (BRCA1) is crucial for DNA damage response (DDR).
• BRCA1 C-terminal (BRCT) domains recognize phosphopeptides containing the pS-X-X-F motif, essential for DDR protein interactions.
• Proteins like ATR-interacting protein (ATRIP) and BRCA1-associated protein required for ATM activation-1 (BAAT1) interact with BRCA1 via this motif.

Purpose of the Study:

• To determine the crystal structures of BRCA1 BRCT domains bound to ATRIP and BAAT1 phosphopeptides.
• To investigate the impact of specific amino acid residues within the phosphopeptide on BRCA1 BRCT binding affinity and specificity.
• To elucidate the molecular mechanisms governing BRCA1's target selection in DNA damage response.

Main Methods:

• X-ray crystallography to determine the structures of BRCA1 BRCTs complexed with ATRIP and BAAT1 phosphopeptides at high resolution (1.75 Å and 2.2 Å).
• Isothermal titration calorimetry (ITC) to quantitatively assess the binding affinities between BRCA1 BRCTs and various phosphopeptides.
• Analysis of peptide-BRCA1 interactions, focusing on the roles of phosphoserine, Phenylalanine at position +3, and residues at positions +1 and +2.

Main Results:

• Crystal structures reveal how pSer and Phe(+3) anchor peptides in the BRCT binding groove.
• The BRCA1-ATRIP structure shows accommodation of Gln(+2) via conformational changes in BRCA1.
• ITC experiments demonstrate that side chains at peptide positions +1 and +2 significantly affect binding affinity, with charged residues like Asp(+1) and Glu(+2) potentially abrogating interaction due to electrostatic repulsion.

Conclusions:

• BRCA1 BRCT domain binding specificity is not solely determined by the canonical pS-X-X-F motif.
• Amino acid residues flanking the core motif, particularly at positions +1 and +2, play a critical role in modulating binding affinity and dictating target selection.
• A systematic evaluation of these flanking residues is necessary to fully understand the molecular basis of BRCA1's function in DDR and its implications in tumorigenesis.