Biophysical and biochemical studies on glycoxidatively modified human low density lipoprotein

Methylglyoxal (MGO), a reactive dicarbonyl metabolite is a potent arginine directed glycating agent which has implications for diabetes-related complications. Dicarbonyl metabolites are produced endogenously and in a state of misbalance, they contribute to cell and tissue dysfunction through protein and DNA modifications causing dicarbonyl stress. MGO is detoxified by glyoxalase 1 (GLO1) system in the cytoplasm. Reactive oxygen species (ROS) are known to aggravate the glycation process. Both the processes are closely linked, and their combined activity is often referred to as "glycoxidation" process. Glycoxidation of proteins has several consequences such as type 2 diabetes mellitus (T2DM), aging etc. In this study, we have investigated the glycation of low-density lipoprotein (LDL) using different concentrations of MGO for varied incubation time periods. The structural perturbations induced in LDL were analyzed by UV-Vis, fluorescence, circular dichroism spectroscopy, molecular docking studies, polyacrylamide gel electrophoresis, FTIR, thermal denaturation studies, Thioflavin T assay and isothermal titration calorimetry. The ketoamine moieties, carbonyl content and HMF content were quantitated in native and glycated LDL. Simulation studies were also done to see the effect of MGO on the secondary structure of the protein. We report structural perturbations, increased carbonyl content, ketoamine moieties and HMF content in glycated LDL as compared to native analog (native LDL). We report the structural perturbations in LDL upon modification with MGO which could obstruct its normal physiological functions and hence contribute to disease pathogenesis and associated complications.