Ligand binding and pH influence conformational flexibility of domains I and III in bovine serum albumin.

Abstract

Bovine Serum Albumin (BSA) is a multi-functional protein that changes its conformation in response to pH and has a high binding capacity for various ligands. Due to these properties, BSA can be used in immunoassay and cell culture applications. To understand how BSA may affect assay function, it is important to understand the flexibility and dynamics of BSA and how it can influence these applications. BSA is a 66-kDa protein composed of three structurally homologous domains. Ligand binding sites can be affected by change in pH as it nears the isoelectric point 4.7-5.2. In our study, we focused on how pH and caprylate impact conformational flexibility of BSA. Using molecular dynamics (MD), we performed simulations using Amber on BSA at pH 7.0 and pH 5.2 both ligand-free and caprylate-bound. We compared all four structures using the root mean square deviation and observed that both the ligand-free and caprylate-bound at pH 7.0 had more structural variation over time than at pH 5.2. The root mean square fluctuation was calculated to compare the conformational flexibility between the residues. We found there were significant conformational differences in Domain I and Domain III at pH 7.0 than pH 5.2 for both ligand-free and caprylate-bound BSA. The surface charge in Domain I is significantly affected with change in pH that may impact the interactions with Domain III and alter binding sites. BSA is a dynamic protein that was found to be structurally different using MD. By understanding BSA conformational flexibility, we can have better insight on its functionality in immunoassay applications.