Cholesterol Content Regulates the Interaction of αA-, αB-, and α-Crystallin with the Model of Human Lens-Lipid Membranes

alpha-Crystallin (alpha ABc) is a major protein comprised of alpha A-crystallin (alpha Ac) and alpha B-crystallin (alpha Bc) that is found in the human eye lens and works as a molecular chaperone by preventing the aggregation of proteins and providing tolerance to stress. However, with age and cataract formation, the concentration of alpha ABc in the eye lens cytoplasm decreases, with a corresponding increase in the membrane-bound alpha ABc. This study uses the electron paramagnetic resonance (EPR) spin-labeling method to investigate the role of cholesterol (Chol) and Chol bilayer domains (CBDs) in the binding of alpha Ac, alpha Bc, and alpha ABc to the Chol/model of human lens-lipid (Chol/MHLL) membranes. The maximum percentage of membrane surface occupied (MMSO) by alpha Ac, alpha Bc, and alpha ABc to Chol/MHLL membranes at a mixing ratio of 0 followed the trends: MMSO (alpha Ac) > MMSO (alpha Bc) approximate to MMSO (alpha ABc), indicating that a higher amount of alpha Ac binds to these membranes compared to alpha Bc and alpha ABc. However, with an increase in the Chol concentration in the Chol/MHLL membranes, the MMSO by alpha Ac, alpha Bc, and alpha ABc decreases until it is completely diminished at a mixing ratio of 1.5. The K-a of alpha Ac, alpha Bc, and alpha ABc to Chol/MHLL membranes at a mixing ratio of 0 followed the trend: K-a (alpha Bc) approximate to K-a (alpha ABc) > K-a (alpha Ac), but it was close to zero with the diminished binding at a Chol/MHLL mixing ratio of 1.5. The mobility near the membrane headgroup regions decreased with alpha Ac, alpha Bc, and alpha ABc binding, and the Chol antagonized the capacity of the alpha Ac, alpha Bc, and alpha ABc to decrease mobility near the headgroup regions. No significant change in membrane order near the headgroup regions was observed, with an increase in alpha Ac, alpha Bc, and alpha ABc concentrations. Our results show that alpha Ac, alpha Bc, and alpha ABc bind differently with Chol/MHLL membranes at mixing ratios of 0 and 0.5, decreasing the mobility and increasing hydrophobicity near the membrane headgroup region, likely forming the hydrophobic barrier for the passage of polar and ionic molecules, including antioxidants (glutathione), creating an oxidative environment inside the lens, leading to the development of cataracts. However, all binding was completely diminished at a mixing ratio of 1.5, indicating that high Chol and CBDs inhibit the binding of alpha Ac, alpha Bc, and alpha ABc to membranes, preventing the formation of hydrophobic barriers and likely protecting against cataract formation.