CLONING/SEQUENCING/EXPRESSION OF HUMAN BSA LIPASE CDNA

人 BSA 脂肪酶 CDNA 的克隆/测序/表达

• 批准号: 3495706
• 负责人: Frederick S Hagen
• 金额: $ 5万
• 依托单位: ZYMOGENETICS, INC.
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-08-01 至 1988-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3495706
• 关键词: Saccharomyces; affinity chromatography; baby food; biotechnology; complementary DNA; developmental nutrition; dietary supplements; enzyme linked immunosorbent assay; enzyme structure; enzyme substrate; gene expression; genetic library; genetic recombination; human milk; hydrolase; lipase; lipolysis; messenger RNA; microorganism mass culture; molecular cloning; nutrition related tag; peptide chemical synthesis; tissue /cell culture

Bile salt-activated lipase (BSAL) is found in the milk of a limited number of mammals: humans, gorillas, dogs, and cats. The enzyme plays an important role in the infant diet in the digestion of lipids and, as such, effects a striking quantitative difference between breast-fed and formula-fed infants in weight gain; in this regard, it is important to note that BSAL is absent in bovine milk. In Phase IB research, we plan to clone, sequence and express the cDNA for human BSAL and provide a framework for Phase II research wherein recombinant human BSAL will be produced in large quantities for supplementation in infant formulas. Expression of the human BSAL cDNA will be achieved by cloning into an expression vector suitable for production at high levels in Saccharomyces cerevisiae. Levels of expression will be ascertained by ELISA and direct measurements of enzymatic activity. The recombinant protein will be purified and characterized for its physical and chemical similarity to the purified human milk enzyme.

胆盐激活脂肪酶（BSAL）存在于有限的 哺乳动物的数量：人类、大猩猩、狗和猫。 的 酶在婴儿饮食中起着重要的消化作用 因此，影响了显著的数量差异， 母乳喂养和配方奶粉喂养的婴儿之间的体重增加;在这个 在这方面，重要的是要注意，BSAL是不存在于牛乳中。 在IB期研究中，我们计划克隆、测序和表达 人BSAL的cDNA，并为II期研究提供框架 研究，其中重组人BSAL将在 大量添加到婴儿配方奶粉中。 人BSAL cDNA的表达将通过克隆 转化到适合于高水平生产的表达载体中， 酿酒酵母 表达水平将是 通过ELISA和直接测量酶 活动 重组蛋白将被纯化， 其特征在于其物理和化学性质与 人乳酶。