Enzymatic deglycation of nonenzymatic glycation products

超出所提供的空间。 糖尿病并发症仍然是糖尿病的一个严重问题。DCCT和UKPDS研究清楚地表明，血糖控制是决定并发症发生率的关键变量。然而，高血糖症与长期并发症的联系机制仍然知之甚少。关于糖尿病并发症的病因学的一个有希望的假说是非酶糖化假说，该假说假定糖尿病相关的病理变化至少部分是葡萄糖及其反应性副产物对蛋白质和一些其它大分子的非酶修饰的结果。该过程中主要的初始中间体是葡萄糖赖氨酸（GL）和果糖赖氨酸（FL）（也称为早期糖化产物- EGP），其经历一系列进一步的重排和反应以形成不可逆的晚期糖化终产物（AGE）。 在我们过去的研究中，我们已经获得证据表明，这种非酶糖化过程是抵消在哺乳动物细胞的主动去糖化机制。基于这些证据，我们建议将糖尿病并发症的非酶糖化假说修改为非酶糖化/酶去糖化假说。我们的第一个假设是，在恒温生物中，蛋白质和其他一些大分子的非酶糖化是不可避免的和有害的。因此，我们的第二个假设是，这些生物体通过主动去糖基化系统控制非酶糖基化的不可食用和破坏性影响。 根据我们的假设，在糖尿病中，特别是在极端高血糖的周期性发作期间，例如在餐后期间观察到的r，非酶糖化过程不受抑制地进行，并且随着时间的推移，导致对必需蛋白质的累积和不可逆的损伤，最终导致细胞功能和活力的丧失。我们目前的研究主要集中在两个方面：1）通过在适当的模型系统中过表达、敲低其mRNA和敲除其基因来检测FN 3 K的功能; 2）研究FN 3 K非依赖性的去糖基化机制。对于FN 3 K的研究，我们已经起草了一份R 01赠款提案给NIDDK（DK 62315 -01）。 为了进行去糖基化的高风险研究，我们提交了这项R21资助提案，我们建议研究转糖基化反应，其中葡萄糖胺的葡萄糖部分（如葡萄糖赖氨酸）被更亲核的化合物（如还原型谷胱甘肽）清除。我们已经有证据表明，这种反应在体外很容易发生，我们也有数据表明，这一过程的主要稳定副产物，葡萄糖-半胱氨酸，存在于尿液中，并在糖尿病患者中升高。这项资助将使我们能够进一步研究转糖基化反应途径，并确定其在血糖正常和糖尿病患者中的重要性。

EXCEED THE SPACE PROVIDED. Diabetic complications remain a serious issue in diabetes mellitus. The DCCT and UKPDS studies demonstrate clearly that glycemic control is a key variable in determining the rate of development of complications. However, mechanisms linking hyperglycemia with long-term complications remain poorly understood. One promising hypothesis on the etiology of diabetic complications is the nonenzymatic glycation hypothesis which postulates that diabetes- associated pathological changes are a result, at least in part, of the nonenzymatic modification of proteins and some other macromolecules by glucose and its reactive byproducts. The predominant initial intermediates in this process are glucoselysine (GL) and fructoselysine (FL) (a.k.a Early Glycation Products - EGP's) which undergo a series of further rearrangements and reactions to form the irreversible Advanced Glycation Endproducts (AGE's). In our past studies we have obtained evidence that this nonenzymatic glycation process is counteractedin mammalian cells by active deglycation mechanisms. Based on this evidence, we are proposing to modify the nonenzymatic glycation hypothesis of diabetic complications to a Nonenzymatic Glycation/Enzymatic Deglycation hypothesis. Our first postulate is that, in homeothermic organisms nonenzymatic glycation of proteins and some other macromolecules is unavoidable and deleterious. Consequently, our second postulate is that these organisms control the unavodable and damaging effects of nonenzymatic glycation by active deglycation systems. According to our hypothesis, in diabetes, especially during periodic episodes of extreme hyperglycemia, such as r observed in the postprandial period, the process of nonenzymatic glycation proceeds unchecked and results, over time, in cumulative and irreversible damage to essential proteins leading ultimately to loss of cell function and viability. Our present research is focused in two areas: 1) Testing the function of FN3K by its overexpression, knockdown of its mRNA and knockout of its gene in appropriate model systems and 2) Study of the FN3K-independent deglycation mechanisms. For the study of FN3K we have drafted a R01 grant proposal to NIDDK (DK62315-01). In order to pursue the higher-risk studies on deglycation we are submitting this R21 grant proposal in which we propose to investigate transglycosylation reactions in which the glucose moiety of glucosamine such as glucoselysine is scavenged away by a more nucleophilic compounds such a reduced glutathione. We already have evidence that such reactions occur readily in vitro and we also have data that the principal stable byproduct of this process, glucose- cysteine, is present in urine and is elevated in diabetic individuals. This grant will enable us to further study the transglycation reaction pathway and to determine its importance in normoglycemic and diabetic individuals PERFORMANCE SITE ========================================Section End===========================================