Multispectral methods combined with computer simulation to explore the binding mechanism of the azo pigment allure red to hemoglobin and myoglobin

Abstract

Allure Red (AR) is a standard food-grade azo pigment and is used in meat products, but the mode of action between allure red and meat-derived proteins is rarely reported. This paper selected two essential proteins in meat, hemoglobin (Hb) and myoglobin (Mb), as protein models to investigate their binding mechanisms with AR. The binding mechanism of AR to Hb/Mb and its conformational changes were investigated using multispectral and computer simulation experiments. The results show that the AR-Hb/Mb system is promoted to bind by hydrogen bonding and van der Waals force; the two systems have a static quenching mechanism at a single binding site. The binding constants (K_a) of the AR-Hb/Mb system are 2.59 × 10⁻⁴ L/mol and 1.05 × 10⁻⁴ L/mol at 277 K. The addition of AR has less effect on the secondary structure of Hb/Mb; the two systems still maintained the structure dominated by α-helix. Computer simulations also showed that the protein system remains stable with the addition of AR, but the Solvent Accessible Surface Area (SASA) increased; the trend was small and did not affect the overall stability of the protein. The study's results help provide theoretical references for the application of AR in related meat products.