AIM: To investigate whether potassium cyanate can inactivate glyceraldehydes 3-phosphate dehydrogenase (GAPDH) and thioltransferase (TTase) in bovine lens.METHODS: Fresh intact bovine lenses were incubated with 100mmol/L potassium cyanate (KCNO) for 7 and 12 days respectively. Then all lens were incubated in 50mmol/L DMEM solution. The proteins in the watersoluble fractions from the normal control and the cyanate-modified lens were extracted. The activity of GAPDH and TTase in the water-soluble fraction after incubation at 37℃ was measured by spectrophotometer.RESULTS: GAPDH activity was significantly lower in the cyanate-modified lens proteins than that of the normal control (P<0.01), and considerably diminished in protein incubated with 100mmol/L potassium cyanate for 12 days. There were statistically significant differences in the activity of TTase between the normal control lenses and the carbamylated lenses incubated for 7 days (P<0.05) and 12 days (P<0.01). However. there was no statistical difference between the samples incubated with 100mmol/L KCNO for 7 and 12 days (P=0.19296).CONCLUSION: This study provides evidence to show carbamylation is able to inactivate GAPDH and TTase in bovine lenses. This may have implications for the susceptibility of lenticular GAPDH and TTase to carbamylation, and also for the research on pathogenesis of cataract.%目的:3-磷酸甘油醛脱氢酶(GAPDH)是糖酵解过程中的一个关键酶,与老化和白内障发生密切相关,硫醇转移酶(TTase)可断裂晶状体蛋白质氧化形成的二硫键使硫醇化的蛋白质脱硫醇,对于维持晶状体透明性极为重要.氨甲酰化为重要的翻译后修饰.本研究观察氨甲酰化是否能诱导晶状体GAPDH和TTase的失活.方法:透明新鲜的晶状体取自2周岁健康牛,与100mmol/L氰酸钾于37℃水孵箱分别振动温育7d和12d.设透明晶状体为正常对照组.所有晶状体孵育在50mmol/L DMEM液中,然后每只晶状体去囊膜,匀浆,提取水溶性蛋白质.用分光光度计分别测量GAPDH和TTase的活性.结果:与氰酸钾孵育后GAPDH的活性与正常对照组相比显著降低(P<0.01),其中12d组与7d组相比,活性明显下降.TTase的活性在孵育7d后与对照组相比有统计学差异(P<0.05),12d组的活性与对照组相比有显著差异(P<0.01),7d组活性与12d组相比无统计学差异(P=0.19296).结论:研究结果显示,分别与100mmol/L氰酸钾孵育7d和12d后,氨甲酰化组的GAPDH和TTase活性与对照组相比有统计学差异,证实氨甲酰化可以诱导GAPDH和TTase酶的失活,为TTase和GAPDH在白内障发病机制中的进一步研究提供了新思路.
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