Structural dynamics of CaMKII activation

描述（由申请人提供）：钙（Ca2+）的浓度增加，可以说是细胞信号传导中最常用的第二信使，激活无处不在的Ca2+传感器钙调蛋白，钙调蛋白随后结合并激活许多酶，包括Ca2+/钙调蛋白依赖性蛋白激酶II （CaMKIl）。CaMKIl作为细胞内钙瞬态的振幅、持续时间和频率所传递的信息的解释器。钙信号的读出是由一系列复杂的构象变化介导的，这些构象变化发生在独特的十二聚体CaMKIl低聚物的范围内。通过自磷酸化，钙浓度恢复到基础水平后，激活持续良好。激活的构象记忆对酶的生理功能至关重要，对学习和记忆是不可或缺的。激活和构象记忆的潜在结构和动态基础尚不清楚。虽然单体催化结构的x射线结构最近已经得到解决，但它的一些特征似乎与生化数据不相容。这项研究的长期目标是通过绘制明确定义的CaMKIl催化中间体的构象变化来弥合结构差距。具体的目标将验证失活CaMKIl被广泛的催化调节相互作用自抑制的假设，Ca2+/CaM结合和自磷酸化通过脱离调节和催化结构域破坏自抑制。我们将利用定向自旋标记和电子顺磁共振（SDSL-EPR）来推导局部环境约束和全局距离约束。我们的初步结果证明了这种方法的可行性，它挑战了单体晶体结构的许多特征。SDSL-EPR研究将扩展到全酶，以获得对协同钙调素结合和自磷酸化自我调节基础的新见解。CaMKIl在学习和记忆中起着至关重要的作用，也可能作为一种自动调节的结构支架，在关键的细胞内位置组装蛋白质复合物，如神经元突触后密度或心脏t小管膜。它与包括帕金森病和精神分裂症在内的许多疾病有关，使其成为药物开发的一个有吸引力的靶点。确定CaMKIl的机制具有基本的生化重要性，可能有助于开发具有更好治疗作用和最小副作用的新化合物。

DESCRIPTION (provided by applicant): Increase in concentration of Calcium (Ca2+), arguably the most utilized second messenger in cellular signaling, activates the ubiquitous Ca2+sensor calmodulin which then proceeds to bind and activate numerous enzymes including Ca2+/calmodulin-dependent protein kinase II (CaMKIl). CaMKIl functions as an interpreter of information conveyed by the amplitude, duration and frequency of intracellular calcium transients. The readout of the calcium signal is mediated by a complex series of conformational changes that occur in the confines of the unique dodecameric CaMKIl oligomer. Activation persists well after return of calcium concentration to basal levels through autophosphorylation. The conformational memory of activation is essential for the enzyme's physiological function and is integral for learning and memory. The underlying structural and dynamic basis of activation and of conformational memory is poorly defined. Although an X-ray structure of a monomeric catalytic domain has recently been solved, some of its features seem incompatible with biochemical data. The long term goal of this research is to bridge the structural gap by mapping conformational changes in well defined catalytic intermediates of CaMKIl. The specific aims will test the hypothesis that inactive CaMKIl is autoinhibited by extensive catalytic-regulatory interaction and that Ca2+/CaM binding and autophosphorylation disrupt autoinhibition by disengaging regulatory and catalytic domains. We will utilize site directed spin labeling and electron paramagnetic resonance (SDSL-EPR) to derive local environmental constraints and global distance restraints. The feasibility of this approach is established in our preliminary results which challenge many features of the monomer crystal structure. The SDSL-EPR studies will be extended to the holoenzyme to gain novel insight into the basis of cooperative calmodulin binding and self regulation by autophosphorylation. CaMKIl plays a critical role in learning and memory and may also serve as an autoregulated structural scaffold to assemble protein complexes at key intracellular locations, such as neuronal postsynaptic densities or on cardiac T-tubule membranes. It is implicated in a number of diseases including Parkinson's disease and Schizophrenia making it an attractive target for drug development. Determining CaMKIl mechanisms is of fundamental biochemical importance and may well facilitate development of new compounds with better therapeutic action and minimal side effects.