STRUCTURAL STUDIES OF CALCIUM/CALMODULIN DEPENDENT KINASE II AND E COLI REPLICA

钙/钙调蛋白依赖性激酶 II 和大肠杆菌复制品的结构研究

• 批准号: 7370608
• 负责人: JOHN KURIYAN
• 金额: $ 0.09万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2007-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7370608
• 关键词: STRUCTURAL; STUDIES; CALCIUM; CALMODULIN; DEPENDENT

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This research is concerned with two classes of protein complexes in which crystals of key assemblies have been obtained. (1) The Calcium/Calmodulin dependent kinase II (CaMKII) is a very large (640 kDa) kinase assembly of 12 monomers known to play an important role in learning and memory. It is normally held in a fully inactive state, with all 12 of its kinases showing only minimal activity. Exposure to calcium bound calmodulin releases the kinase activity, however the details of the molecular mechanism of this process have been impossible to induce from biochemical studies. Understanding the regulatory mechanism of the kinase will require a detailed molecular model of both active and inactivate forms of the kinase. Recently, we have obtained crystals from the C. elegans CamKII orthologue. Initial diffraction experiments performed at the ALS indicate that these crystals diffract x-rays isotropically to ~7.5 ¿¿ and weakly to 3.9 ¿¿in certain directions. Preliminary analysis of the unit cell and non-crystallographic symmetry elements indicates that the crystals contain an intact holoenzyme of 12 CaMKII molecules (about 640kDa) in the unit cell, however the data is very weak and its quality is not sufficient to allow for further analysis. (2)DNA Replication. (i) DNA polymerase is the enzyme responsible for replicating chromosomes. The structures of several polymerases from various organisms have been determined, however, the structure of the major E. coli replicative DNA polymerase has thus far remained elusive. Recently, we obtained crystals of the E. coli polymerase catalytic subunit. The crystals are small (~50 ¿¿m), and diffract x-rays weakly to ~4¿¿.

该子项目是利用 NIH/NCRR 资助的中心拨款提供的资源的众多研究子项目之一，该子项目和研究者 (PI) 可能已从其他 NIH 来源获得主要资助，因此可以在其他 CRISP 机构中得到体现。列出的是该中心，该中心不一定是研究者的机构。本研究涉及两类蛋白质复合物，其中已获得关键组件的晶体。 (1) 钙/钙调蛋白依赖性激酶 II。 (CaMKII) 是一种非常大的 (640 kDa) 激酶组合，由 12 个单体组成，已知在学习和记忆中发挥重要作用。它通常处于完全非活性状态，其所有 12 个激酶仅表现出最低限度的活性。钙结合的钙调蛋白释放激酶活性，然而，从生化研究中不可能得出该过程的分子机制的细节，了解激酶的调节机制将需要活性和失活形式的详细分子模型。最近，我们从线虫 CamKII 直系同源物中获得了晶体，在 ALS 上进行的初步衍射实验表明，这些晶体的 X 射线衍射各向同性约为 7.5 ¿ ¿并弱至 3.9 ¿ ¿对晶胞和非晶体学对称元素的初步分析表明，晶体中含有12个CaMKII分子（约640kDa）的完整全酶，但数据非常薄弱，其质量不足以证明。 (2)DNA 复制 (i) DN​​A 聚合酶是负责复制染色体的酶，但来自不同生物体的几种聚合酶的结构已被确定。迄今为止，主要的大肠杆菌复制 DNA 聚合酶的结构仍然难以捉摸。最近，我们获得了大肠杆菌聚合酶催化亚基的晶体。这些晶体很小（约 50 微米），X 射线衍射能力较弱。至~4¿ ¿ 。