Cholesterol and Q147E Deamidation Modulates αA-Crystallin Membrane Binding Elucidating Protective Role of Lens...

The αA-Crystallin (αAc) binding with lens membranes increases with age and cataract formation. However, the role of lipids and cholesterol (Chol) in Q147E-αAc membrane binding remains unclear, which we aim to elucidate in this study.

We have used the electron paramagnetic resonance spin-labeling method to probe the Chol/ 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) and Chol/ sphingomyelin (SM) membranes binding with wild-type (WT) and Q147E-αAc.

Compared to WT-αAc, the Q147E mutant had increased binding to POPC and decreased binding to SM membranes without Chol. Adding 33 mol% Chol to the POPC and SM membranes decreased the binding of WT and, to a lesser degree, decreased the binding of Q147E-αAc to the membranes. Adding 60 mol% Chol completely inhibited Q147E mutant and WT binding to POPC membranes. However, 33 and 60 mol% Chol completely inhibited WT and Q147E mutant binding to SM membranes, respectively. WT and Q147E-αAc membrane binding decreased membrane mobility while increasing order and hydrophobicity near the headgroup.

In Chol-free membranes, the deamidated Q147E-αAc binds significantly more to the POPC membranes compared to WT, whereas WT binds significantly more to the SM membranes compared to Q147E-αAc. In contrast, for 33 mol% Chol-containing membranes, the deamidated Q147E-αAc binds significantly more to POPC and SM membranes than WT. Conversely, 60 mol% Chol-containing membranes completely inhibit WT and deamidated Q147E-αAc binding to POPC and SM membranes. These results suggest that increased Chol content of the lens membranes during aging protects against accumulation of modified proteins on the membrane associated with cataracts.