Structural basis for a hand-like site in the calcium sensor CatchER with fast kinetics.
Structural basis for a hand-like site in the calcium sensor CatchER with fast kinetics.
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DOI：
10.1107/s0907444913021306
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发表时间：
2013-12
期刊：
影响因子：
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通讯作者：
Ying Zhang;Florence N. Reddish;Shen Tang;You Zhuo;Yuan‐Fang Wang;Jenny J. Yang;I. Weber
中科院分区：
文献类型：
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作者：
Ying Zhang;Florence N. Reddish;Shen Tang;You Zhuo;Yuan‐Fang Wang;Jenny J. Yang;I. Weber
钙离子是一种重要的信号分子，通过绿色荧光蛋白（GFP）的工程突变体CatchER可以快速检测到内质网中的钙离子。高分辨率（1.78-1.20 Å）分析了载脂蛋白形式和与钙或钆配合物的CatchER晶体结构，以探测金属离子的结合位点。虽然CatchER在溶液中表现出1:1的结合化学计量，但在突变残基S147E、S202D、Q204E、F223E和T225E的羧酸侧链形成的手状位点上，观察到每种金属离子的结合位置都有两个，这可能是其快速动力学性质的原因。通过对CatchER、野生型GFP和增强型GFP的结构比较，证实了Thr203和Glu222的不同构象与发色团的Tyr66的两种形式有关，这两种构象负责不同蛋白质的吸收波长。钙与CatchER的结合改变了Glu222侧链构象居群的平衡，进一步改变了其光学性质。
Calcium ions, which are important signaling molecules, can be detected in the endoplasmic reticulum by an engineered mutant of green fluorescent protein (GFP) designated CatchER with a fast off-rate. High resolution (1.78-1.20 Å) crystal structures were analyzed for CatchER in the apo form and in complexes with calcium or gadolinium to probe the binding site for metal ions. While CatchER exhibits a 1:1 binding stoichiometry in solution, two positions were observed for each of the metal ions bound within the hand-like site formed by the carboxylate side chains of the mutated residues S147E, S202D, Q204E, F223E and T225E that may be responsible for its fast kinetic properties. Comparison of the structures of CatchER, wild-type GFP and enhanced GFP confirmed that different conformations of Thr203 and Glu222 are associated with the two forms of Tyr66 of the chromophore which are responsible for the absorbance wavelengths of the different proteins. Calcium binding to CatchER may shift the equilibrium for conformational population of the Glu222 side chain and lead to further changes in its optical properties.