Enhanced degradation and decreased stability of eye lens alpha-crystallin upon methylglyoxal modification.
Methylglyoxal (MGO), a potent glycating agent, forms advanced glycation end products (AGEs) with proteins. Several diabetic complications including cataract are thought to be the result of accumulation of these protein-AGEs. alpha-Crystallin, molecular chaperone of the eye lens, plays an important role in maintaining the transparency of the lens by preventing the aggregation/inactivation of several proteins/enzymes in addition to its structural role. Binding of adenosine-5-triphosphate (ATP) to alpha-crystallin has been shown to enhance its chaperone-like function and protection against proteolytic degradation. In the earlier study, we have shown that modification of alpha-crystallin by MGO caused altered chaperone-like activity along with structural changes, cross-linking, coloration and subsequent insolubilization leading to scattering of light [Biochem. J. 379 (2004) 273]. In the present study, we have investigated ATP binding, stability and degradation of MGO-modified alpha-crystallin. Proteolytic digestion with trypsin and chymotrypsin showed that MGO-modified alpha-crystallin is more susceptible to degradation compared to native alpha-crystallin. Furthermore, ATP was able to protect native alpha-crystallin against proteolytic cleavage but not MGO-modified alpha-crystallin. Interestingly, binding studies indicate decreased ATP binding to MGO-modified alpha-crystallin and support the decreased protection by ATP against proteolysis. In addition, differential scanning calorimetric and denaturant-induced unfolding studies indicate that modification of alpha-crystallin by MGO leads to decreased stability. These results indicate that MGO-modification of alpha-crystallin causes partial unfolding and decreased stability leading to enhanced proteolysis. Cross-linking of these degraded products could result in aggregation and subsequent insolubilization as observed in senile and diabetic cataract lenses.
Satish Kumar
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M
Department of Biochemistry, National Institute of Nutrition (ICMR), Jamai-Osmania, Hyderabad 500 007, India.
Mrudula
T
T
Mitra
N
N
Bhanuprakash Reddy
G
G
D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D002417:Cattle; D002918:Chymotrypsin / Q000737:chemistry; D007908:Lens, Crystalline / Q000737:chemistry*; D018832:Molecular Chaperones / Q000737:chemistry; D017510:Protein Folding; D011765:Pyruvaldehyde / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000737:chemistry; D038201:alpha-Crystallins / Q000737:chemistry* / Q000187:drug effects
D018832:Molecular Chaperones; D038201:alpha-Crystallins; D011765:Pyruvaldehyde; D000255:Adenosine Triphosphate; D002918:Chymotrypsin; D014357:Trypsin