CRYOELECTRON MICROSCOPY OF ACTOS1 ATPASE INTERMEDIATES

ACTOS1 ATP酶中间体的冷冻电子显微镜

• 批准号: 2910889
• 负责人: HOWARD D. WHITE
• 金额: $ 36.52万
• 依托单位: EASTERN VIRGINIA MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-04-01 至 2004-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2910889
• 关键词: actins; adenosine diphosphate; bioimaging /biomedical imaging; chemical kinetics; cryoelectron microscopy; enzyme mechanism; enzyme structure; enzyme substrate complex; freezing; hydrolysis; image processing; muscle contraction; myosins; nucleotide analog; site directed mutagenesis; stop flow technique

DESCRIPTION (Adapted from abstract): The molecular basis of muscle force is a configurational change in the myosin head while attached to actin. Such a change could either be a conformational change within the head itself, or it may be an alteration in the binding angle made with actin. The energy for force production is known to come from ATP hydrolysis, but while the "tilting cross-bridge" hypothesis remains attractive, it is still unproven. Moreover, recent structural data have suggested that the configuration of attached myosin heads may be similar in both the ATP and ADP-Pi states. One problem in testing the tilting bridge hypothesis has been the difficulty of establishing the structures of so-called weakly bound acto-myosin states that occur in the presence of ATP. A considerable amount is known about the structure of the strongly bound states that occur in ADP or no nucleotide. However, other bound states have been elusive either because they dissociate at the low protein concentrations required for moderate resolution microscopy of the acto-myosin head (S1) complex or they are present only transiently. The applicant recently found conditions that allow one to observe transient and weakly bound states and has been studying their structures by electron cryo-microscopy of hydrated specimens. The micrographs already obtained show a variety of attached S1 configurations during steady-state ATP hydrolysis; such data are therefore compatible with the tilting bridge hypothesis but do not prove it.

描述（改编自摘要）：肌肉力量的分子基础是 附着在肌动蛋白上时肌球蛋白头部的构型变化。这样的 变化可能是头部本身的构象变化，或者 可能是与肌动蛋白结合角度的改变。力的能量 已知生产来自ATP水解，但虽然“倾斜 “跨桥”假说仍然很有吸引力，但它仍然未经证实。此外，委员会认为， 最近的结构数据表明，附着的肌球蛋白的构型 在ATP和ADP-Pi状态下，头部可能相似。 在测试倾斜桥假说的一个问题是， 建立所谓的弱结合肌动蛋白-肌球蛋白的结构， 在ATP存在下发生。有相当多的人知道 在ADP或无核苷酸中发生的强结合态的结构。 然而，其他的束缚态一直难以捉摸，因为它们解离在 中等分辨率显微镜所需的低蛋白浓度 肌动蛋白-肌球蛋白头（S1）复合物或它们仅短暂存在。的 申请人最近发现了允许人们观察瞬态和 弱束缚态，并一直在研究它们的结构， 水合标本的冷冻显微镜检查。已经获得的显微照片显示， 在稳态ATP水解过程中各种附着的S1构型;例如 因此，数据与倾斜桥假设是一致的，但不 证明给我看