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REGULATION OF BROWN ADIPOSE TISSUE THERMOGENESIS

棕色脂肪组织产热的调节

基本信息

批准号：
3231068
负责人：
EARL S SHRAGO
金额：
$ 7.29万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
1983
资助国家：
美国
起止时间：
1983-08-01 至 1992-07-31
项目状态：
已结题

项目摘要

The hypothesis that nonshivering thermogenesis (NST) mediated by brown adipose tissue mitochondria (BATM) is linked to obesity and the regulation of body weight has been recently proposed. The molecular basis for this weight regulating ability is the occurrence of a nucleotide binding protein called uncoupling protein (UP) within the BATM inner membrane which imparts the unique capability of controlled uncoupling of respiration from normal rgulatory mechanisms thus permitting loss of energy as heat. The metabolic regulation of the UP and thus NST has not been ascertained. It is known that purine nucleotides will bind and inhibit the function of the UP. However, no activator of the UP function has been clearly established. We have recently purified and reconstituted the UP into a liposome system and observed faty acid activation of proton transport. In future studies we will attempt to purify the protein to homogeneity for use in the reconstituted system to determine its kinetic properties and mechanism of action by fatty acids. The UP will be extracted with Triton X-100 from mitochondria, purifed on hydroxylapatite and the Triton exchanged with octylglucoside for final reconstitution into a liposome system. Proton conductance will be assayed using the pH system as well as incorporation of cytochrome oxidase with the UP. Measurements of O2 consumption which would be more physiological could then be carried out. The mechanism of fatty acid activation of the UP will be studied using spin labeled fatty acids and measurement by electron spin resonance. Individual intracellular free fatty acids and their long chain thioesters will be determined by HPLC and thier concentrations related to activation of the UP. Hamsters will be the main source of BATM for purification and biochemical studies relating to mechanism of the UP. However, more physiological experiments will be carried out by reconstituting the UP from cold adapted rats and the ob/ob mouse which will provide the information as to whether the UP is non functional under certain conditions or merely decreased in concentration. The long term objectives of the proposal are to determine the mechanism of action of the UP in BATM and how certain aspects of the biochemical and molecular action of the protein might be extended to understanding basal energy expenditure in humans.

棕色介导的非颤抖产热（NST）假说脂肪组织线粒体（BATM）与肥胖及其调节有关最近有人提出了体重的影响。其分子基础体重调节能力是核苷酸结合蛋白的出现 BATM 内膜内称为解偶联蛋白 (UP)，它赋予控制呼吸与正常情况脱钩的独特能力调节机制允许能量以热量的形式损失。新陈代谢 UP 的监管以及 NST 尚未确定。我们都知道嘌呤核苷酸会结合并抑制 UP 的功能。然而，尚未明确确定 UP 功能的激活剂。我们最近将 UP 纯化并重构为脂质体系统，观察到脂肪酸激活质子运输。在今后的研究中我们将尝试将蛋白质纯化至均质以用于重构系统以确定其动力学特性和机制脂肪酸的作用。 UP 将使用 Triton X-100 从线粒体，在羟基磷灰石上纯化，并与 Triton 交换辛基葡萄糖苷最终重构为脂质体系统。质子电导率将使用 pH 系统以及掺入细胞色素氧化酶与 UP。测量 O2 消耗量然后可以进行更加生理化的治疗。脂肪的作用机制将使用自旋标记脂肪酸研究 UP 的酸活化和通过电子自旋共振测量。个体细胞内游离脂肪酸及其长链硫酯将通过 HPLC 测定它们的浓度与 UP 的激活有关。仓鼠将是 BATM 的主要来源，用于纯化和生化研究 UP机制。然而，更多的生理学实验将会通过从冷适应大鼠和 ob/ob 中重建 UP 来进行鼠标将提供有关 UP 是否为非的信息在某些条件下起作用或只是浓度降低。该提案的长期目标是确定以下机制： UP 在 BATM 中的作用以及生化和代谢的某些方面如何蛋白质的分子作用可能会扩展到理解基础人体的能量消耗。

项目成果

期刊论文数量（3）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Reconstitution of purified brown adipose tissue mitochondria uncoupling protein: demonstration of separate identity of nucleotide binding and proton translocation sites by chemical probes.

纯化的棕色脂肪组织线粒体解偶联蛋白的重建：通过化学探针证明核苷酸结合和质子易位位点的单独身份。

DOI：
10.1073/pnas.86.8.2559
发表时间：
1989
期刊：
Proceedings of the National Academy of Sciences of the United States of America
影响因子：
11.1
作者：
Katiyar,SS;Shrago,E
通讯作者：
Shrago,E

Characterization of a low molecular weight protein of the ATP synthetase complex from beef heart and rat liver mitochondria with a high affinity monoclonal antibody.

使用高亲和力单克隆抗体对来自牛心和大鼠肝线粒体的 ATP 合成酶复合物的低分子量蛋白质进行表征。