Biomolecular Interactions of RAD51181–200 with BRCA1846–871 and Mutants and Molecular Docking Approach

Abstract

Affecting the repair function of RAD51 can effectively reverse tumor resistance to chemotherapy and radiotherapy. There is interaction between RAD51181−200 and BRCA1846−871. Using the computer to simulate single mutation of lysine at 862 position of BRCA1846−871, six BRCA1846−871 mutants with significant interaction with RAD51181−200 were screened out. These mutant peptides were obtained by 9-fluorenylmethyloxycarbonyl (Fmoc) solid-phase peptide synthesis (SPPS) method. All peptides were purified and characterized by liquid chromatography and ESI-MS. In this study, the interaction between RAD51181−200 and BRCA1846−871 or mutant peptides was investigated using circular dichroism spectroscopy (CD) and fluorescence spectroscopy. The CD results shown that the secondary structure change of RAD51181−200 induced by BRCA1846−871, BRCA1846−871 K862F (F), BRCA1846−871 K862G (G), BRCA1846−871 K862E (E) and BRCA1846−871 K862N (N) was obvious, which the α-helix content of RAD51181−200 decreased from 58.3–36.1%, 58.3–36.2%, 58.3–36.4%, 58.3–36.5% and 58.3–36.0%, respectively. While, the addition of BRCA1846−871 K862P (P) and BRCA1846−871 K862Y (Y) hardly changed the α-helix content of RAD51181−200. From the fluorescence results, the binding constants of F and Y are 1.00 × 105 M−1 and 1.16 × 105 M−1 respectively, which were both larger than that of BRCA1846−871 (9.01 × 102 M−1). The binding constant of RAD51181−200 with E (7.45 × 103 M−1) is slightly higher than that of BRCA1846−871. The binding constants of RAD51181−200 with P (1.24 × 103 M−1) and N (1.26 × 103 M−1) are similar to that of BRCA1846−871. However, the Kb value of G (1.51 × 102 M−1) is less than that of BRCA1846−871. Comparing all spectral data, F is the most significant one interacting with RAD51181−200. These findings could provide direction for peptide processing and application.