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BIOPHYSICS AND ENZYMOLOGY OF KINESIN MOVEMENT

驱动蛋白运动的生物物理学和酶学

基本信息

批准号：
6385968
负责人：
JEFF GELLES
金额：
$ 29.79万
依托单位：
BRANDEIS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1991
资助国家：
美国
起止时间：
1991-04-01 至 2002-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6385968
关键词：
biomechanics cell motility chemical conjugate conformation enzyme activity enzyme structure intracellular transport kinesin microtubules nanotechnology protein structure function site directed mutagenesis

项目摘要

Kinesin is a mechanoenzyme that drives microtubule-based intracellular organelle transport processes. Kinesin couples a free-energy-liberating chemical reaction (the hydrolysis of ATP) to a cycle of mechanical processes that move the enzyme molecules and attached organelles along microtubules. We want to characterize the cycle of mechanical processes by which kinesin moves and to determine how these processes are coupled to the reactions of ATP hydrolysis. We have developed novel experimental systems that allow us to directly monitor nanometer-scale mechanical processes, domain movements, and chemical steps in single isolated kinesin molecules using light microscope-based instruments. Intracellular organelle transport by kinesin and kinesin homologs plays an essential role in the physiology of eukaryotic cells. Its functions include transport of materials, chromosome and nuclear movements in mitosis/meiosis, and morphogenesis of membranous organelles. To explore these functions at the molecular level we will: Characterize the conformational changes in the kinsin alpha-helical coiled-coil neck domain that are required for processive motility along microtubules. Test the hand-over-hand hypothesis for the kinesin movement mechanism Detect and characterize the power stroke of an isolated kinesin head domain. The experiments will combine single-molecule biophysics techniques with conventional enzymology methods including protein chemical modification, site-directed mutagenesis, and chemical kinetics techniques.

动蛋白是一种机械酶，可以驱动基于微管的细胞内细胞器运输过程。Kinesin配对释放自由能量化学反应(三磷酸腺苷的水解)形成一个机械循环移动酶分子和附着的细胞器的过程微管。我们想要描述机械过程的循环并确定这些过程是如何耦合的三磷酸腺苷的水解反应。我们已经开发出了小说允许我们直接监测纳米级的实验系统机械过程、磁区移动和化学步骤使用光学显微镜仪器分离动蛋白分子。动蛋白及其同系物在细胞器内的转运作用在真核细胞的生理学中起着重要作用。它的功能包括物质的运输、染色体和核的运动有丝分裂/减数分裂和膜细胞器的形态发生。去探索这些在分子水平上的功能我们将：表征激动素α-螺旋的构象变化前向运动所需的卷曲颈区微管。检验Kinesin运动机制的手-手假说检测和表征孤立运动蛋白头部的力量冲程域。这些实验将结合单分子生物物理技术和传统的酶学方法包括蛋白质化学修饰，定点突变和化学动力学技术。

项目成果

期刊论文数量（19）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Single molecule approaches for studying spliceosome assembly and catalysis.

研究剪接体组装和催化的单分子方法。

DOI：
10.1007/978-1-62703-980-2_17
发表时间：
2014
期刊：
Methods in molecular biology (Clifton, N.J.)
影响因子：
0
作者：
Anderson,EricG;Hoskins,AaronA
通讯作者：
Hoskins,AaronA

Three-color single molecule imaging shows WASP detachment from Arp2/3 complex triggers actin filament branch formation.

DOI：
10.7554/elife.01008
发表时间：
2013-09-03
期刊：
eLife
影响因子：
7.7
作者：
Smith BA;Padrick SB;Doolittle LK;Daugherty-Clarke K;Corrêa IR Jr;Xu MQ;Goode BL;Rosen MK;Gelles J
通讯作者：
Gelles J

Mechanism of transcription initiation at an activator-dependent promoter defined by single-molecule observation.

DOI：
10.1016/j.cell.2012.01.018
发表时间：
2012-02-17
期刊：
Cell
影响因子：
64.5
作者：
Friedman LJ;Gelles J
通讯作者：
Gelles J

Ordered and dynamic assembly of single spliceosomes.

DOI：
10.1126/science.1198830
发表时间：
2011-03-11
期刊：
Science (New York, N.Y.)
影响因子：
0
作者：
Hoskins AA;Friedman LJ;Gallagher SS;Crawford DJ;Anderson EG;Wombacher R;Ramirez N;Cornish VW;Gelles J;Moore MJ
通讯作者：
Moore MJ

Cease-fire at the leading edge: new perspectives on actin filament branching, debranching, and cross-linking.

DOI：
10.1002/cm.20543
发表时间：
2011-11
期刊：
CYTOSKELETON
影响因子：
2.9
作者：
Ydenberg, Casey A.;Smith, Benjamin A.;Breitsprecher, Dennis;Gelles, Jeff;Goode, Bruce L.
通讯作者：
Goode, Bruce L.