KINETICS OF B12, INTRINSIC FACTOR, AND OTHER PROTEINS

该项目的长期目标是准确描述 钴胺素与肠道所需大分子的相互作用 这种维生素的吸收，运输和细胞摄取，并发展 更容易应用的放射性测定，通过其来识别、测量和 研究生理性和非生理性钴的代谢。 为了实现这些目标，我们有以下具体目标： 表征内在因子（IF）-钴胺素 复合物与IF受体蛋白结合， 钴胺素的内化。 为此，我们将使用放射免疫测定法， 回肠上皮细胞的免疫细胞化学分析。 B）识别和 表征钴胺素结合蛋白的免疫反应形式 通过凝胶过滤和离子交换色谱分离， 特异性RIA来鉴定这些蛋白质。 c）发展放射免疫分析 测量钴胺素的特定生理形式的技术及其应用 这些测定辅酶浓度的分析方法 直接在血液和组织提取物中。 d）分离和表征 用色谱法测定血浆中非生理形式的类咕啉化合物 分离和配体结合放射性测定。 （e）发展 放射免疫酶测定研究两种生理 辅酶，腺苷钴胺素和甲基钴胺素，并表征 这些生物合成过程所需的酶。 本项目中获得的信息和开发的技术将 扩展我们对钴胺素吸收障碍的理解， 可能与药物、毒素和肝脏疾病一起发生的转运;以及 测量钴胺素辅酶的分析技术可以提供一个远 确定获得性和遗传性细胞疾病的简单方法 钴胺素辅酶合成。

The long term objectives of this project are to characterize precisely the interaction of cobalamin with macromolecules required for intestinal absorption, transport, and cellular uptake of this vitamin and to develop more readily applicable radioassays by which to identify, measure, and study the metabolism of the physiologic and non-physiologic cobalamins. Toward these objectives, we have the following specific aims: a) To characterize the mechanism by which the intrinsic factor (IF)-cobalamin complex binds to the IF-receptor protein and facilitates the internalization of cobalamin. For this we will use radioimmunoassay and immunocytochemical analysis of ileal epithelial cells. b) To identify and characterize immunoreactive forms of the cobalamin binding proteins fractionated by gel-filtration and ion exchange chromatography using specific RIA to identify these proteins. c) To develop radioimmunoassay techniques to measure the specific physiologic forms of cobalamin and use these analytical methods to measure the concentration of these coenzymes directly in blood and tissue extracts. d) To isolate and characterize the nonphysiologic forms of corrinoid compounds in plasma using chromatographic separation and ligand binding radioassays. e) To develop radioimmunoenzymatic assays to study the synthesis of the two physiologic coenzymes, adenosyl cobalamin and methyl cobalamin and to characterize the enzymes required for these biosynthetic processes. The information obtained and the techniques developed in this project would extend our understanding of the disorders of cobalamin absorption and transport which may occur with drugs, toxins and liver disease; and the analytical techniques to measure cobalamin coenzymes could provide a far simpler method for determining acquired and inherited cellular disorders of cobalamin coenzyme synthesis.