COOPERATIVE EFFECTS IN SMOOTH MUSCLE REGULATION

平滑肌调节的协同作用

• 批准号: 2439644
• 负责人: SHERWIN LEHRER
• 金额: $ 17.06万
• 依托单位: BOSTON BIOMEDICAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-05-15 至 1998-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2439644
• 关键词: actins; analytical ultracentrifugation; animal tissue; calcium flux; caldesmon; calponin; chemical kinetics; conformation; enzyme activity; fluorescence microscopy; fluorescent dye /probe; microfilaments; muscle contraction; myosins; phosphorylation; protein isoforms; protein structure function; smooth muscle; striated muscles; tropomyosin; troponin

Despite the importance of smooth muscle in health and disease, the details of its operation and regulation remain largely unclear. This renewal application is a continued effort intended to gain more information in this area. The overall goal is to use molecular and cellular methodologies to gain information on the functional roles of various smooth muscle proteins and to enhance our understanding of the regulatory mechanism of smooth muscle contraction. The program contains six projects, two facility cores and an administrative core. Projects I- A deals with the effect of phosphorylation of the myosin structure and on the interaction between acting and myosin. Project I-B is concerned with the mechanism by which myosin light chain kinase is activated by Ca/calmodulin and that by which myosin light chain is recognized by the enzyme. Projects II-A and II-B study the structure and function of caldesmon and calponin, respectively, in the hope that such studies will lead to the elucidation of their roles in the thin filament-based regulation of smooth muscle contraction. Project II-C searches for the regulatory pathway involving the mitogen activated protein kinases in smooth muscle. In Project III the integrated system of the thick and the thin filaments will be examined, and the possible role of tropomyosin in the cooperative properties of smooth muscle thin filament will be explored. All projects, with the exception of II-C and II, use chemical crosslinking and resonance energy transfer as tools to characterize protein-protein interactions, and practically every project uses-directed mutagenesis to produce protein variants. Projects II-A II-C, in particular, will explore genetic approaches to elucidate the functional significance of some of the regulatory proteins. Other methods include fluorescence and circular dichroism measurements, chemical and enzymatic proteolysis, use of synthetic peptides, analytical ultracentrifugation and electron microscopy. The Biophysical/Biochemical Core includes Protein Chemistry service, the Analytical Ultracentrifugation service and the Electron Microscopy and Immunocytochemistry service. In the Protein Expression ore, recombinant proteins will be generated by baculovirus expression system in insect cells. Both cores will be used by all projects. Funding of this program will allow us to continue our investigation on the regulatory mechanisms in the smooth muscle system.

尽管平滑肌对健康和疾病很重要， 其运作和监管的细节在很大程度上仍不清楚。这 续签申请是一项持续的努力，旨在获得更多 该领域的信息。总体目标是利用分子和 细胞方法学来获取有关功能作用的信息 各种平滑肌蛋白并增强我们对 平滑肌收缩的调节机制。 该计划 包含六个项目、两个设施核心和一个行政核心。 项目 I-A 涉及肌球蛋白磷酸化的影响 结构以及作用和肌球蛋白之间的相互作用。 项目 I-B 涉及肌球蛋白轻链的机制 激酶由 Ca/钙调蛋白和肌球蛋白光激活 链被酶识别。 项目 II-A 和 II-B 研究 caldesmon 和 calponin 的结构和功能分别 希望此类研究能够阐明它们在 基于细丝的平滑肌收缩调节。 项目 II-C 寻找涉及的监管途径 平滑肌中有丝分裂原激活蛋白激酶。 在项目三中 粗丝和细丝的集成系统将是 检查，以及原肌球蛋白在合作社中的可能作用 将探索平滑肌细丝的特性。 全部 除 II-C 和 II 之外的项目均使用化学交联 和共振能量转移作为表征蛋白质-蛋白质的工具 相互作用，几乎每个项目都使用定向诱变 产生蛋白质变体。 特别是项目 II-A II-C 探索遗传学方法来阐明其功能意义 一些调节蛋白。 其他方法包括荧光 和圆二色性测量，化学和酶法 蛋白水解、合成肽的使用、分析超速离心 和电子显微镜。 生物物理/生化核心包括 蛋白质化学服务、分析超速离心服务 以及电子显微镜和免疫细胞化学服务。 在 蛋白质表达矿石，重组蛋白质将通过以下方式产生 昆虫细胞中的杆状病毒表达系统。 两个核心都会被使用 由所有项目。 该计划的资金将使我们能够继续 我们对平滑肌调节机制的研究 系统。