GLYCOLIPID AND GLYCOPROTEIN METABOLISM IN EYE TISSUE

眼组织中的糖脂和糖蛋白代谢

• 批准号: 2157903
• 负责人: EDWARD L KEAN
• 金额: $ 33.41万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-04-01 至 1997-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2157903
• 关键词: Golgi apparatus; RNase protection assay; carbohydrate structure; chick embryo; cow; dolichol; electron microscopy; enzyme activity; galactosyltransferases; glycolipids; glycoprotein biosynthesis; immunoaffinity chromatography; immunocytochemistry; isozymes; laboratory mouse; laboratory rat; light microscopy; lipid metabolism; messenger RNA; oligosaccharides; recombinant DNA; retina; retinoid binding proteins; rhodopsin; rod cell

The oligosaccharide chains of bovine rhodopsin are unique when compared to other glycoproteins in that they are abridged in size and are hybrids of high mannose and complex types. We are investigating mechanisms used by the retina that give rise to this situation, i.e., what controls are present such that galactose and additional GlcNAc residues, that would be foci for oligosaccharide chain extension reactions, are not found (except in trace amounts) in mature rhodopsin. We shall extend our previous investigations on the kinetics of the galactosyltransferases that demonstrated the low efficiency of the retina enzyme toward intact rhodopsin and opsin. We now will use oligosaccharides and glycopeptides prepared from rhodopsin to explore the influence of the protein matrix on these reactions. A reference compound for these studies will be the interphotoreceptor retinoid binding protein (IRBP), a naturally galactosylated glycoprotein of the retina. Galactosyltransferases purified from bovine retina, from rat liver Golgi, and from milk will be examined. We shall investigate by immunocytochemistry the localization of galactosyltransferase in the photoreceptor cell using both light and electron microscopy. Using the techniques of molecular biology we shall investigate whether there are differences in the relative amounts of the mRNAs that code for the long and short forms of galactosyltransferase in the retina and the photoreceptor cell. Are these different isoforms differentially located in the photoreceptor cell; is the isoform found in the retina concerned mainly with surface recognition in contrast to biosynthetic reactions? Investigations will be carried out concerning control by the retina of GlcNAc-transferase II, an enzyme that could add a GlcNAc to the alpha(1 - > 6) mannose arm of the rhodopsin oligosaccharides and thus be a site for chain extension. The kinetics of this reaction, using rhodopsin, opsin, rhodopsin oligosaccharides and glycopeptides as acceptors, will be examined using the enzyme obtained by recombinant DNA techniques. We shall also examine for the presence in the retina of galactosidase and hexosaminidase activity toward pertinent analogs of rhodopsin. We shall attempt the isolation of bovine photoreceptor cells by immunoaffinity chromatography. The structure of the oligosaccharides of human rhodopsin will be investigated. We shall continue our investigations concerning control of the initiating reactions of the dolichol pathway in the retina. We shall examine the influence on the allosteric activation by dolichol-P-mannose of GlcNAc- lipid synthesis (a phenomenon which we have discovered) by other key intermediates of the pathway. The findings of mutations in the rhodopsin gene (some in sites controlling glycosylation) in patients with retinitis pigmentosa have highlighted the importance of understanding the chemistry and biochemistry of the glycosylation of rhodopsin. The investigations concerning control of the dolichol pathway may further our basic understanding of the regulation of glycoprotein biosynthesis.

相比之下，牛视紫红质的低聚糖链是独特的