Effect of local anesthetic drug procaine hydrochloride on the conformational stability of bovine hemoglobin: Multi-spectroscopic and computational approaches.

The interaction between bovine hemoglobin (BHb) and local anesthetic drug procaine hydrochloride (PCH) was examined by spectroscopic and computational studies. Intrinsic fluorescence analysis explored the ground-state complex formation in the binding of PCH with BHb through static quenching mechanism. The binding constants () are 29.38 × 10, 22.54 × 10 and 17.99 × 10 at 288, 298 and 308 K, respectively, and the ratio of BHb:PCH was 1:1 in the interaction mechanism of PCH and BHb. The acquired thermodynamic parameters ( and ) demonstrated that interaction mechanism is spontaneous and enthalpy driven. The van der Waals forces and hydrogen bonding have been played a predominant role in the binding mechanism. The UV-vis spectroscopy validates the ground-state complexation between PCH and BHb and the binding constant () has been evaluated utilizing Benesi-Hildebrand equation. Fluorescence resonance energy transfer (FRET) results have demonstrated that the distance between donor (BHb) and acceptor (PCH) is very short (2.34 nm) suggesting a significant probability to energy transfer from BHb to PCH. Synchronous fluorescence results revealed that the alteration in the micro-environment of Tyrosine (Tyr) is more than tryptophan (Trp) residues suggesting that PCH molecule is close to Tyr residue. The secondary structure alterations were confirmed by CD, 3-D fluorescence and FT-IR spectroscopic measurements. Moreover, computational analyses further corroborated that PCH molecules are closer to Tyr residues as compared to Trp residues of BHb during the interaction process. The BHb-PCH complexes may contribute to a deeper understanding of the metabolism of drug, blood circulation process and may help to illustrate the relationship between functions and structure of BHb.Communicated by Ramaswamy H. Sarma.