Temperature and cholesterol regulate the association of native bovine lens alpha-crystallin with in vitro reconstituted membranes made of synthetically prepared lipids and cholesterol.

α-Crystallin (αABc) association with the lens membrane is accelerated in cataracts, and cholesterol (Chol) in lens membranes is accelerated in healthy aging and decreases with cataracts. This study investigates how temperature and Chol affect the membrane interactions of αABc. We applied the electron spin resonance method to analyze the membrane association of αABc incubated with (Chol)/model of bovine lens-lipid (MBLL) membranes at temperatures of 30, 37, 45, and 52 °C with Chol/MBLL mixing ratios of 0.0, 0.3, and 1.5. The highest percent of membrane exterior surface occupied (HMEO) by αABc to Chol/MBLL membranes at mixing ratios of 0.0 and 0.3 resulted in the tendency: (37 °C) HMEO > (45 °C) HMEO ≈ (52 °C) HMEO > (30 °C) HMEO. Additionally, the binding affinity (Ka) of αABc to Chol/MBLL membranes at mixing ratios of 0 and 0.3 resulted in the trends (37 °C) Ka > (30 °C) Ka > (45 °C) Ka ≈ (52 °C) Ka. This data indicates that incubation temperature regulates the total amount bound and the affinity of αABc to Chol/MBLL membranes. However, with all investigated temperatures, the HMEO and Ka of αABc to Chol/MBLL membrane were zero at a Chol/MBLL ratio of 1.5, implying that high Chol and cholesterol bilayer domains (CBDs) formation totally inhibits αABc membrane association independent of temperature. Furthermore, our data show that temperatures regulate the hydrophobicity and mobility near the surface of Chol/MBLL membranes at a mixing ratio of 0.0 and 0.3 with αABc association. In contrast, at a mixing ratio of 1.5, no significant changes in hydrophobicity and mobility were observed. This study showed that the HMEO by αABc and Ka of αABc to membranes are differentially modulated by temperature, altering hydrophobicity and mobility near the membrane surface. However, Chol and CBDs reduce the membrane association of αABc irrespective of temperature, ultimately helping to maintain the lens function, transparency, homeostasis, and preventing cataract development.