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Molecular Mechanisms of Protein Crosslinking in the Lens

晶状体中蛋白质交联的分子机制

基本信息

批准号：
8482333
负责人：
Ram H Nagaraj
金额：
$ 39.63万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2017-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8482333
关键词：
Advanced Glycosylation End Products Aging Animal Model Arginine Ascorbic Acid Binding Cataract Cell Nucleus Cells Chemicals Crystallins Data Deamination Detection Development Environment Etiology Eye Goals Human In Vitro Incubated Innovative Therapy Kynurenine Laboratories Lead Lysine Mass Spectrum Analysis Measures Mediating Mediator of activation protein Methods Modification Molecular Nuclear Organ Culture Techniques Oxygen Oxygen measurement, partial pressure, arterial Paper Pathway interactions Pattern Physiological Pigmentation physiologic function Pigments Play Post-Translational Protein Processing Process Prodrugs Protein Binding Proteins Reaction Reduced Glutathione Risk Factors Role Seminal Senile Cataract Structure Surface Testing Transgenic Animals Transgenic Mice Tryptophan Tryptophan 2,3 Dioxygenase Ultraviolet A radiation Water Work adduct aged analytical method ascorbate base crosslink efficacy testing inhibitor/antagonist interest lens lens cortex lens protein molecular mass novel oxidation pentosidine prevent protein aggregate protein aggregation protein crosslink public health relevance research study

项目摘要

Our long-term goal is to prevent human cataracts by understanding the molecular mechanisms involved. This project builds on our previous work of nearly 25 years on lens protein modifications in aging and cataractogenesis. Protein crosslinking is a major modification in aged and cataractous lenses. Ascorbate (ASC) is a major constituent of the lens, which in the human lens is present at concentrations up to 2 mM. ASC is oxidized in aged and cataractous lenses, and its oxidation products react rapidly with lens proteins to form pigmented and crosslinked proteins through formation of advanced glycation end products (AGEs). Reduced glutathione (GSH) offers some protection against this process, but the decreased levels of GSH in aged and cataractous lenses favor ASC oxidation. Recent work suggests that much of the protein crosslinking in cataractous lenses are ASC oxidation product-mediated. We know that ASC is oxidized in aging and cataractous lenses, but we do not know the mechanisms for such oxidation. Although molecular oxygen- mediated oxidation is likely to occur in the cortex, it is unlikely to occur in the near anoxic nucleus. Despite this limitation, protein crosslinking and aggregation through AGE formation is most prominent in the nucleus of cataractous lenses. Kynurenines are tryptophan oxidation products produced by the kynurenine pathway initiated by indoleamine 2,3-dioxygenase. They are present in relatively high levels in human lenses. Kynurenines undergo spontaneous deamination and bind covalently to lens proteins. Our preliminary studies show that both protein-free and protein-bound kynurenines promote ASC oxidation. UVA light has been considered as an important risk factor for cataractogenesis, although the mechanisms are still obscure. Our preliminary experiments suggest that kynurenine-mediated ASC oxidation is significantly accelerated by UVA light, and that such oxidation can occur both in the presence and absence of oxygen. Based on these observations, we hypothesize that kynurenine-mediated ASC oxidation followed by protein modification plays an important role in the etiology of senile cataracts. We will test this hypothesis with the following three aims. In aim 1 we will determine kynurenine-mediated ASC oxidation in the presence and absence of oxygen and UVA light, conditions that emulate cortex and nucleus of the human lens. In aim 2 we will determine the impact of kynurenine-mediated ASC oxidation on covalent crosslinking and aggregation of lens proteins, and in aim 3, we will test our newly developed prodrug compounds on Kyn/ASC-mediated protein modification and crosslinking, and evaluate their effects on cataract development. Together, the proposed studies will unravel the interplay between kynurenines and ASC in lens protein modification in human cataracts, and the findings could lead to innovative therapies to prevent or delay cataracts in humans.

我们的长期目标是通过了解所涉及的分子机制来预防人类白内障。这