PROTEINASE INHIBITORS & CRYSTALLIN FRAGMENTS IN CATARACT

描述：蛋白质的修饰在将正常透镜转变为白内障透镜中起关键作用。内源性酶对晶体蛋白中的肽键的蛋白水解切割可能是最常见和最重要的修饰之一。到目前为止，关于晶体蛋白中这些切割的信息在其酶性质、在酶和底物水平的调节、对晶体蛋白功能的影响以及晶体蛋白片段中的二级修饰方面是有限的。根据初步结果，P.I.假设晶体蛋白的降解在酶和底物水平上都受到调节，并且切割的片段经历二次修饰，例如通过Maillard反应的糖化，因此直接参与透镜蛋白的交联，这是透镜混浊发展的核心。为了检验这一假设，将追求以下三个具体目标：（1）具体目标#1：确定膜蛋白酶的自溶激活机制，该机制来自去污剂如脱氧胆酸钠对BA 31 A1-晶体蛋白酶的推定酶原型激活。(2)具体目标#2：确定膜蛋白酶和BA 31 A1-晶状体蛋白酶在选定晶状体蛋白体内蛋白水解中的作用。(3)具体目标#3：确定降解多肽的交联，特别是9 kDa yD-晶状体蛋白片段的交联，以形成同源（仅晶状体蛋白片段）和异源（晶状体蛋白片段加天然晶状体蛋白）多聚体，可能通过美拉德反应的糖化机制。上述研究将解决有关晶体蛋白降解的体内调节和内源性蛋白酶在这一过程中的作用的问题。这项研究还将回答的问题是否降解的多肽，对二级修饰，获得交联性能和直接参与蛋白质交联过程中的透镜混浊的发展。由于所有拟议的研究都将在人类晶状体中进行，因此结果将与人类老年性白内障问题直接相关。

DESCRIPTION: Modification of proteins play a pivotal role in changing a normal lens into a cataractous lens. Proteolytic cleavage of peptide bonds in crystallins by endogenous enzymes is probably one of the most frequent and important modifications. So far, information about these cleavages in crystallins is limited in terms of their enzymatic nature, regulation at the enzyme and substrate levels, effect on crystallin functions and secondary modifications in crystallin fragments. Based on preliminary results, the P.I. has hypothesized that the degradation of crystallins is regulated at both enzyme and substrate levels and the cleaved fragments undergo secondary modification such as glycation via the Mail lard reaction and therefore directly participate in the cross-linking of the lens proteins which is central to the development of lens opacity. To test this hypothesis, the following three specific aims will be pursued: (1) specific Aim #1: Determine the mechanism of autolytic activation of a membrane proteinase from a putative zymogen type activation of the BA31A1-crystallin proteinase by a detergent such as sodium deoxycholate. (2) specific Aim #2: Determine the role of the membrane proteinase and BA31A1-crystallin proteinase in the in vivo proteolysis of selected crystallins. (3) Specific Aim #3: Determine crosslinking of degraded polypeptides in general and specifically of a 9 kDa yD-crystallin fragment to form homologous (crystallin fragments alone) and heterologous (crystallin fragments plus native crystallins) multimers possibly via the glycation mechanism of the Maillard reaction. The above studies will address the questions regarding the in vivo regulation of crystallin degradation and the role of endogenous proteinases in this process. This study will also answer the questions whether the degraded polypeptides, on secondary modifications, acquire cross-linking properties and participate directly in protein cross-linking during the development of lens opacity. Since all of the proposed studies will be carried out in human lenses, the results will be directly relevant to the human senile cataract problem.