Abstract
Aims/hypothesis Hyperglycaemia, one of the main features of diabetes, results in non-enzymatic glycation of plasma proteins, including apolipoprotein A-I (apoA-I), the most abundant apolipoprotein in HDL. The aim of this study was to determine how glycation affects the structure of apoA-I and its ability to activate lecithin:cholesterol acyltransferase (LCAT), a key enzyme in reverse cholesterol transport.
Materials and methods Discoidal reconstituted HDL (rHDL) containing phosphatidylcholine and apoA-I ([A-I]rHDL) were prepared by the cholate dialysis method and glycated by incubation with methylglyoxal. Glycation of apoA-I was quantified as the reduction in detectable arginine, lysine and tryptophan residues. Methylglyoxal-AGE adduct formation in apoA-I was assessed by immunoblotting. (A-I)rHDL size and surface charge were determined by non-denaturing gradient gel electrophoresis and agarose gel electrophoresis, respectively. The kinetics of the LCAT reaction was investigated by incubating varying concentrations of discoidal (A-I)rHDL with a constant amount of purified enzyme. The conformation of apoA-I was assessed by surface plasmon resonance.
Results Methylglyoxal-mediated modifications of the arginine, lysine and tryptophan residues in lipid-free and lipid-associated apoA-I were time- and concentration-dependent. These modifications altered the conformation of apoA-I in regions critical for LCAT activation and lipid binding. They also decreased (A-I)rHDL size and surface charge. The rate of LCAT-mediated cholesterol esterification in (A-I)rHDL varied according to the level of apoA-I glycation and progressively decreased as the extent of apoA-I glycation increased.
Conclusions/interpretation It is concluded that glycation of apoA-I may adversely affect reverse cholesterol transport in subjects with diabetes.
Original language | English |
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Journal | Diabetologia |
Volume | 50 |
Issue number | 3 |
Pages (from-to) | 643-653 |
Number of pages | 11 |
ISSN | 0012-186X |
DOIs | |
Publication status | Published - Mar 2007 |
Keywords
- apolipoprotein A-I
- diabetes
- high-density lipoproteins
- lecithin : cholesterol acyltransferase
- non-enzymatic glycation
- HIGH-DENSITY-LIPOPROTEINS
- NONENZYMATIC GLYCOSYLATION
- MICELLAR COMPLEXES
- DIABETES-MELLITUS
- METHYLGLYOXAL
- PROTEINS
- BINDING
- VITRO
- AGE
- HDL