THREE TYPES OF GLYCOSYLTRANSFERASES

三种类型的糖基转移酶

• 批准号: 3172905
• 负责人: KHUSHI L MATTA
• 金额: $ 13.5万
• 依托单位: ROSWELL PARK CANCER INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-07-01 至 1996-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3172905
• 关键词: affinity chromatography; carbohydrate structure; chemical synthesis; enzyme activity; enzyme inhibitors; enzyme mechanism; enzyme structure; enzyme substrate; fucose; galactosyltransferases; glycosides; glycosyltransferase; high performance liquid chromatography; human subject; human tissue; neoplasm /cancer diagnosis; neoplastic cell; oligosaccharides; protein purification; sialyltransferases; tumor antigens

Our primary objective is to identify and characterize new types of glycosyltransferases responsible for biosynthetic pathways which are different or reverse to the currently accepted pathways. We plan to identify alpha(1,2)-L-fucosyltransferase which can incorporate fucose at the C-2 position of galactose in the X-determinant (Le(x)) structure, Galbeta1 yields 4 (Fucalpha1 yields 3) GlcNAc, to give the Le(y) sequence, Fucalpha1 yields 2 Galbeta1 yields 4 (Fucalpha1 yields 3) GlcNAcbeta1 yields OR. Similarly, we plan to study the alpha (2,3)- sialyltransferases which can use Le(x) and Le(a) structures as acceptors. Recently, we have isolated and partially characterized beta(1-3)-galactosyltransferase which incorporates galactose to the alpha-linked GalNAc residue in the R-GlcNAcbeta1 yields 6 GalNAcalpha1 yields OB sequence to give R-GlcNAcbeta1 yields 6 (Galbeta1 yields 3) GalNAcalpha1 yields OR (Core II structure). We are interested in utilizing different tumor cell lines and both normal and cancerous human tissues as source material for our investigations. We will also attempt to purification of some of these new enzyme activities, such as beta(1,3)-galactosyltransferase and alpha(1,3)-L-fucosyltransferase present in human lung carcinoma cell lines. Affinity chromatography will be employed as a key step in these purifications. Both nucleotide and specific-acceptor ligands will be made available for this technique. Our chemical synthetic approach will be primarily used for the procurement of the variety of well-defined carbohydrate structures required for the study of these enzymes, including acceptors and ligands. Certain unique and highly specific acceptor moieties can be provided to enable us to examine the activity of a single glycosyltransferase activity in spite of the presence of other glycosyltransferases present in the same source. We also intend to evaluate some of our modified analogs as inhibitors of certain glycosyltransferase activities. The identification of new types of glycosyltransferase activities will shed more light on the biosynthetic pathways of glycoconjugates, especially fucosylated glycoconjugates containing Le(x), Le(a) and sialyl (Le(x) and Le(a)) determinants. We intend through this research to fill the various gaps that exist in our knowledge of the specificity of those enzymes proposed for study. A successful outcome of the proposed program will open various avenues toward other long-term objectives. For example, the discovery and documentation of glycosyl- transferases of novel specificities can enable the development of unique enzymatic and immunological probes to detect these enzymes.

我们的主要目标是识别和表征新类型的 负责生物合成途径的糖基转移酶， 与目前接受的途径不同或相反。 我们计划 鉴定可掺入岩藻糖α（1，2）-L-岩藻糖基转移酶 在X-决定簇中半乳糖的C-2位（Le（x）） 结构，Galbeta 1产生4（Fucalpha 1产生3）GlcNAc，得到 Le（y）序列，Fucalpha 1产生2 Galbeta 1产生4（Fucalpha 1产生 3)GlcNAc β 1产生OR。 同样，我们计划研究alpha（2，3）- 可以使用Le（x）和Le（a）结构的唾液酸转移酶， 受体。 最近，我们分离并部分鉴定了 β（1-3）-半乳糖基转移酶，其将半乳糖掺入到 R-GlcNAc β 1中的α-连接的GalNAc残基产生6个GalNAcalpha 1 产生OB序列，得到R-GlcNAc β 1产生6（Galbeta 1产生3） GalNAcalpha 1产生OR（核心II结构）。 我们有兴趣 利用不同的肿瘤细胞系， 人体组织作为我们研究的原始材料 我们还将 尝试纯化这些新酶活性中的一些，例如 作为β（1，3）-半乳糖基转移酶和α（1，3）-L-岩藻糖基转移酶 存在于人肺癌细胞系中。 亲和层析 将被用作这些纯化的关键步骤。 两个核苷酸 和特异性受体配体将可用于此 法 我们的化学合成方法将主要用于 获得各种明确的碳水化合物结构 这些酶的研究所需的，包括受体和 配体。 某些独特的和高度特异性的受体部分可以是 提供使我们能够检查单个活动的活动 糖基转移酶活性，尽管存在其他 糖基转移酶存在于同一来源。 我们还打算 评估我们的一些修饰类似物作为某些 糖基转移酶活性。 新型糖基转移酶活性的鉴定将 揭示了糖缀合物的生物合成途径， 特别是含有Le（x）、Le（a）和 sialyl（Le（x）和Le（a））决定簇。 我们打算通过这项研究 来填补我们对这些特殊性的认识上的各种空白 这些酶的研究。 取得圆满成果 拟议的计划将开辟各种途径，以其他长期 目标. 例如，糖基的发现和记录- 新特异性的转移酶可以使开发 独特的酶和免疫探针来检测这些酶。