摘要
目的鉴定早期糖基化晶状体蛋白质。方法新鲜牛晶状体匀浆与[14C]果糖孵育24h,凝胶过滤层析(SephacrylS300HR)分离水溶性蛋白质,并进一步分离脱氢酶;采用亲合层析(Affigel601)分离糖基化蛋白质;SDS聚丙烯酰胺凝胶电泳分离和鉴定富含放射活性组份的性质。结果α、β晶状体蛋白和某些脱氢酶为早期糖基化蛋白质。再次分离表明αA2和βA3晶状体蛋白为首攻糖基化蛋白质;亲合层析结果显示分子质量约为20kDa的可能为α和γ晶状体蛋白,36kDa的多肽可能为苹果酸盐脱氢酶或乳酸脱氢酶,这些均为早期糖基化蛋白质。结论αA2、βA3、γ晶状体蛋白和某些脱氢酶是最容易被糖基化的晶状体蛋白质。糖基化导致晶状体蛋白质的结构发生改变和蛋白质凝聚。早期糖基化诱导酶(如脱氢酶)的失活,可能参与白内障的形成。
Objective To identify the primary targets of glycation in the lens,Methods The fresh bovine lens homogenate was incubated with[^14C] fructose for 24 hours. The water-soluble proteins were separated by size-exclusion chromatography （Sephacryl S-300HR） and then were passed over an enzyme column to separate out the dehydrogenases.Glycated proteins from the water-soluble fractions were separated by using a sugar affinity column （Affi-Gel 601）.Then the radioactive fractions were identified on SDS gels and further separated and characterised.Results α,βcrystallin and some dehydrogenases showed high levels of early glycatiou products.The further separation revealed that αA2 and βA3-crystallin had preferential glycation.Affinity chromatography results indicated proteins with a subtmit weight of 20 kDa which could correspond to an α and γ-crystallin, along with a 36 kDa possibly corresponding to malate dehydeogenase or lactate dehydro- genase were labelled at an early stage.Conclusion αA2,βA3,γ-crystallin and possibly some dehydrogenase are the most susceptible lens proteins to glycatioa. Glycation of lens proteins could result in its structural alteratiou and subsequently aggregation.The early glycatiou may induce the inactivation of the enzymes （eg.dehydrogenase） and possibly play a role in the pathogenesis of cataract formation. [ Rec Adv Ophthalmol 2005 ; 25 （5） ： 404-407 ]
出处
《眼科新进展》
CAS
2005年第5期404-407,共4页
Recent Advances in Ophthalmology
基金
英国国际发展研究基金资助(编号:AL070667)~~
