CALCIUM BINDING PROTEIN/TARGET INTERACTIONS

许多细胞内的过程是由细胞内的瞬时变化调节的 钙离子浓度。钙离子的作用是由蛋白质介导的 与钙离子特异结合，并呈现新的构象，然后 使它们能够与目标蛋白质结合，包括一些酶，从而 影响它们的功能。这个项目的目的是了解 两者最能表征钙离子结合的分子机制 调节蛋白，钙调蛋白(CaM)和肌钙蛋白C(TNC)，识别和 以依赖于钙离子的方式调节它们的靶标。 钙离子和钙离子诱导CaM和TNC的局部和整体构象变化 通过与选定的靶点-整体蛋白(磷酸二酯酶， 钙调神经磷酸酶、神经调制蛋白、肌钙蛋白I)和模拟 互动网站-将被描述为。人肝细胞钙调素和钙调素的突变体 兔骨骼TNC将由关键位置的半胱氨酸残基合成。 将监测半胱氨酸侧链之间距离的变化 在用适合荧光能量的发色团标记它们之后 转移测量。特定词语的功能意义 CaM和TNC的构象转变将通过评估 突变体的靶标调节能力，其中选择的运动 蛋白质片段受到二硫键的限制。互动网站 带有目标分子的CaM和TNC将通过 旨在定义专一性的决定因素。突变体也将被用来 评估某些疏水基团和离子基团在特定条件下的贡献 位置与相互作用的强度和调节活性有关。 这些研究将有助于更好地了解心力衰竭的发病机制。 CaM、TNC等钙离子结合蛋白的钙信号转导。这个 交互接口和定义的详细说明 特异性的决定因素将是重要的鉴定 快速增长的家庭中其他成员的生理指标 钙结合蛋白。

Many intracellular processes are regulated by a transient change in the concentration of Ca2+ ions. The effect of Ca2+ is mediated by proteins that bind Ca2+ specifically and assume a new conformation, which then enables them to bind to target proteins, including some enzymes, thereby affecting their function. The aim of this project is to understand molecular mechanisms by which the two best characterized Ca2+-binding regulatory proteins, calmodulin (CaM) and troponin C (TnC), recognize and modulate their targets in a Ca2+-dependent way. Local and global conformational changes induced in CaM and TnC by Ca2+ and by interactions with selected targets - whole proteins (phosphodiesterase, calcineurin, neuromodulin, troponin I) and peptides that mimic the interaction sites - will be characterized. Mutants of human liver CaM and rabbit skeletal TnC will by synthesized with Cys residues at key positions. Changes in distances between the Cys side chains will be monitored following their labeling with chromophores suitable for fluorescence energy transfer measurements. The functional significance of specified conformational transitions in CaM and TnC will be assessed by evaluating the target-modulating ability of mutants in which the movement of selected protein segments is restricted by a disulfide bond. Sites of interaction with target molecules on CaM and TnC will be characterized in detail with a view to defining determinants of specificity. Mutants will also be used to assess the contribution of some hydrophobic and ionic groups at specific positions to the strength of interaction and to the modulatory activity. These studies will contribute to a better understanding of the mechanism of Ca2+ signal transduction by CaM, TnC and other Ca2+-binding proteins. The detailed description o the interaction interfaces and definition of determinants of specificity will be important for identification of physiological targets for other members of the rapidly growing family of Ca2+-binding proteins.