Biophysical and computational approaches to unravel the molecular interaction mechanism of bromodeoxyuridine, a proliferative marker with human serum albumin

The interaction of a nucleoside analogue bromodeoxyuridine (BrdU) with human serum albumin (HSA) was studied to investigate the binding phenomenon and analyse the protein conformation upon BrdU binding. Multiple spectroscopic techniques, viz. intrinsic and 3-D fluorescence, UV–Vis absorption, and circular dichroism (CD) spectroscopy along with molecular docking were used. Decrease in the Stern–Volmer constant (K_sv) as well as the association constant (K_a) with increase in temperature suggested BrdU–HSA complex formation. Intermediate binding affinity between BrdU and HSA was evident from the K_a values (2.49–3.97 × 10⁴ mol^–1 dm³), while BrdU–HSA complex formation was driven by hydrophobic and van der Waals interactions along with hydrogen bonds, as revealed by thermodynamic data (ΔS = + 28.48 J mol⁻¹ K⁻¹; ΔH = − 17.16 kJ mol⁻¹). Minor changes occur in both secondary and tertiary structures as well as in the fluorophores' microenvironment of HSA, as recognized from the CD spectral results in the far-UV and the near-UV regions and 3-D fluorescence spectra, respectively. Use of site markers (warfarin and indomethacin for site I; diazepam for site II) as well as docking results suggested BrdU binding to both site I (more preferred) and site II, located in subdomains IIA and IIIA, respectively, of HSA. Graphic abstract:

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