Interaction of alpha-crystallin with four major phospholipids of eye lens membranes

It is well-studied that the significant factor in cataract formation is the association of alpha-crystallin, a major eye lens protein, with the fiber cell plasma membrane of the eye lens. The fiber cell plasma membrane of the eye lens consists of four major phospholipids (PLs), i.e., phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), and sphingomyelin (SM). Despite several attempts to study the interaction of alpha-crystallin with PLs of the eye lens membrane, the role of individual PL for the binding with alpha-crystallin is still unclear. We recently developed the electron paramagnetic resonance (EPR) spin-labeling method to study the binding of alpha-crystallin to the PC membrane (Mainali et al., 2020a). Here, we use the recently developed EPR method to explicitly measure the binding affinity (K-a) of alpha-crystallin to the individual (PE*, PS, and SM) and two-component mixtures (SM/PE, SM/PS, and SM/PC in 70:30 and 50:50 mol%) of PL membranes as well as the physical properties (mobility parameter and maximum splitting) of these membranes upon binding with alpha-crystallin. One of the key findings of this study was that the K-a of alpha-crystallin binding to individual PL membranes followed the trends: K-a(PC) > K-a(SM) > K-a(PS) > K-a(PE*), indicating PE* inhibits binding the most whereas PC inhibits binding the least. Also, the K-a of alpha-crystallin binding to two-component mixtures of PL membranes followed the trends: K-a(SM/PE) > K-a(SM/PS) > K-a(SM/PC), indicating SM/PC inhibits binding the most whereas SM/PE inhibits binding the least. Except for the PE* membrane, for which there was no binding of alpha-crystallin, the mobility parameter for all other membranes decreased with an increase in alpha-crystallin concentration. It represents that the membranes become more immobilized near the headgroup regions of the PLs when more and more alpha-crystallin binds to them. The maximum splitting increased only for the SM and the SM/PE (70:30 mol%) membranes, with an increase in the binding of alpha-crystallin. It represents that the PL headgroup regions of these membranes become more ordered after binding of alpha-crystallin to these membranes. Our results showed that alpha-crystallin binds to PL membranes in a saturable manner. Also, our data suggest that the binding of alpha-crystallin to PL membranes likely occurs through hydrophobic interaction between alpha-crystallin and the hydrophobic fatty acid core of the membranes, and such interaction is modulated by the PL headgroup's size and charge, hydrogen bonding between headgroups, and PL curvature. Thus, this study provides an in-depth understanding of alpha-crystallin interaction with the PL membranes made of individual and two-component mixtures of the four major PLs of the eye lens membranes.

Automated summary

This study focuses on the association of alpha-crystallin, a major eye lens protein, with different phospholipids of the eye lens membrane. By using the electron paramagnetic resonance method, the binding affinity of alpha-crystallin to individual phospholipids and two-component mixtures of phospholipids was measured, revealing varying effects on the binding. The research sheds light on the interaction of alpha-crystallin with phospholipid membranes and provides insights into the factors influencing this interaction.