PROPERTIES OF DYNEIN ISOENZYMES

动力蛋白同工酶的特性

基本信息

批准号：
3278159
负责人：
IAN R GIBBONS
金额：
$ 22.24万
依托单位：
UNIVERSITY OF HAWAII AT MANOA
依托单位国家：
美国
项目类别：
财政年份：
1982
资助国家：
美国
起止时间：
1982-04-01 至 1992-03-31
项目状态：
已结题

项目摘要

The specific aims of this project are to determine the physicochemical and enzymatic properties of the multiple isoenzymic forms of dynein ATPase present in cilia and sperm flagella, to elucidate the steps in the mechanochemical cycle by which the binding and hydrolysis of molecules of ATP at sites on the outer and inner dynein arms generate mechanical shear stresses between adjacent doublet tubules of the axoneme, to clarify the function of two distinct ATPase sites present in the dynein-1 outer arm, and possibly also in the dynein-2 inner arm, and to investigate the extent to which the interaction of the dynein isoenzymes with micromolar concentrations of Ca2+ are involved in the Ca2+-regulation of the form and direction of bending in cilia and sperm flagella. We shall look for partial functional activity in subsets of heavy and light polypeptide chains in the 21S outer arm particle in sea urchin sperm flagella and for a significant interaction with calcium and an intrinsic calmodulin-type regulatory subunit. Efforts will be made to solubilize dynein-2 in a form that retains functional capability as a preliminary to determining its function. Photocleavage of the dynein heavy chains in the presence of vanadate and ATP will be used in conjunction with photoaffinity labelling and monoclonal antibodies to map the functional domains to specific regions of the polypeptides. The interaction of the various dynein isoenzymes with native tubulin and with other axonemal proteins will be studied in order to obtain information about the function of the isoenzymes in flagellar motility. The long-term objective of the project is to elucidate the functional role of the dynein isoenzymes in ciliary and flagellar motility, as well as in the forms of cytoplasmic movement involving cytoplasmic microtubules. This work is of direct relevance to problems of male infertility resulting from sperm immotility as in Kartageners syndrome and to the severe respiratory problems associated with the recently defined "immotile-cilia syndrome".

该项目的具体目的是确定物理化学和多种酶形式的动力蛋白ATPase的酶特性出现在纤毛和精子鞭毛中，以阐明机械化学循环，通过这些循环的结合和水解分子的结合和水解在外部和内动力蛋白臂上的位置ATP产生机械剪切轴突的相邻双重小管之间的应力，以澄清 Dynein-1外臂中存在两个不同ATPase位点的功能，并可能在Dynein-2内臂中，并研究程度动力蛋白同工酶与微摩尔的相互作用 Ca2+的浓度参与了形式的Ca2+调节纤毛和精子鞭毛的弯曲方向。我们将寻找重度多肽子集的部分功能活性海胆精子鞭毛中21秒的链条中的链条和与钙和固有的钙调蛋白类型的显着相互作用监管亚基。将努力以一种形式溶解Dynein-2 保留功能能力作为确定其的初步功能。在存在的情况下钒酸盐和ATP将与光性标签一起使用和单克隆抗体将功能域映射到特定区域多肽。各种动力蛋白同工酶与将研究天然小管蛋白和其他轴突蛋白获取有关鞭毛中同工酶功能的信息运动。该项目的长期目标是阐明动力蛋白同工酶在纤毛和鞭毛运动中的功能作用，以及涉及细胞质的细胞质运动形式微管。这项工作与男性问题直接相关精子免疫产生的不育症，如卡牛蛋白综合征和与最近定义的严重呼吸系统问题 “ Immotile-Cilia综合征”。