Exploring the interaction between ABCL and HAG: a combined experimental and theoretical approach

Abstract

Abemaciclib (ABCL), as cell cycle-dependent kinase 4/6 inhibitor, is used to treat a variety of tumors, including breast, melanoma, and bladder cancer. The purpose of this paper was to comprehend the binding mechanism of ABCL to the plasma transport protein HAG using multi-spectroscopic approaches, molecular docking, and dynamic simulations. The outcomes revealed that the quenching constant of HAG caused by ABCL declined with the temperature rising, indicating that ABCL could extinguish endogenous fluorescence of HAG through the static quenching method. Thermodynamic analyses, competition experiments, and molecular dynamics simulations indicate that the main driving forces between ABCL and HAG are hydrogen bonding, hydrophobic interactions, and Van der Waals forces. The binding distance between the two was calculated from the FRET theory to be 2.94 nm. Molecular docking showed that ABCL binds within the hydrophobic cavity of HAG. Molecular dynamics simulations confirmed that the original configuration of HAG changed slightly upon incorporation of ABCL. Synchronous fluorescence analysis revealed a hypsochromic shift (∼1 nm) in the maximum emission wavelength of Tyr residues. This blue displacement phenomenon correlates with decreased microenvironmental polarity, likely induced by ABCL binding through two distinct mechanisms, including steric shielding effect and conformational restriction. Far-UV CD spectral results demonstrated subtle conformational perturbations in HAG upon ABCL binding, including α-helix content decreased from 1.84 % to 1.67 % and β-sheet content reduced from 41.92 % to 41.85 %. In addition, some common metal ions, for instance, Cu²⁺, Mg²⁺, Fe³⁺, Ca²⁺ and Zn²⁺ may cause the alteration in the binding constant between ABCL and HAG, resulting in changes in the action of ABCL.