Proximity relationship in the binary complex formed between troponin I and troponin C.
Proximity relationship in the binary complex formed between troponin I and troponin C.
复制标题
肌钙蛋白 I 和肌钙蛋白 C 之间形成的二元复合物中的邻近关系。
DOI：
10.1016/0022-2836(86)90145-2
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发表时间：
1986
影响因子：
5.6
通讯作者：
Cheung,HC
中科院分区：
文献类型：
--
作者：
Wang,CK;Cheung,HC
我们已经确定了六个分子之间的距离在肌钙蛋白C（TnC）和肌钙蛋白I（TnI）之间形成的二元复合物的供体和受体探针，无论是内在的荧光团（Trp 158的TnI）或附着到网站的外来探针之间的荧光共振能量转移的四个网站。3种外源探针分别为丹磺氮丙啶（DNZ）、N′-（碘乙酰基）-N′-（8-磺基-1-萘基）乙二胺（IAEDANS）和5-（碘乙酰氨基）曙红（IAE）。四种荧光团提供四种供体-受体对：DNZ→ IAE、Trp→ IAEDANS、IAEDANS→ IAE和Trp→ DNZ。它们允许通过测量从（1）TnC中的Met 25（DNZ）到Cys 98（IAE）的能量转移，（2）TnI中的Trp 158到Cys 133（IAEDANS），（3）TnC的Cys 98（IAEDANS）到TnI的Cys 133（IAE），（4）TnI的Trp 158到TnC的Cys 98（IAEDANS），（6）TnC的Met 25（DNZ）与TnI的Cys 133（IAE）之间的相互作用。当分离的蛋白质与TnI复合时，TnC中的距离（1）几乎不受影响，而当TnI掺入二元复合物中时，TnI中的距离（2）增加了6倍（29%）。在EGTA存在的情况下，基于κ 2= 2 3，复合物中的六个供体-受体分离（R）在28至57 Å范围内。Mg 2+对R的影响很小，而Ca 2+在6个距离中的5个距离上使R显著增加或降低。这些变化并不伴随着显着的变化，在轴向去极化的荧光团。结果表明，钙离子与肌钙蛋白C的结合引起了复合物中两种蛋白质区域的整体结构扰动，提示肌钙蛋白亚基结构域的大规模运动可能是钙离子调节收缩过程中钙离子信号传递的起始分子事件。
We have determined six molecular distances among four sites in the binary complex formed between troponin C (TnC) and troponin I (TnI) by fluorescence resonance energy transfer between donor and acceptor probes that were either an intrinsic fluorophore (Trp158 of TnI) or extrinsic probes attached to the sites. The three extrinsic probes were dansylaziridine (DNZ), N′-(iodoacetyl)-N′-(8-sulfo-1-naphthyl) ethylenediamine (IAEDANS) and 5-(iodoacetamido) eosin (IAE). The four fluorophores provided four donor-acceptor pairs: DNZ→ IAE, Trp→ IAEDANS, IAEDANS→ IAE, and Trp→ DNZ. They allowed determinations of separations between specific sites from measurements of energy transfer from (1) Met25 (DNZ) to Cys98 (IAE) in TnC,(2) Trp158 to Cys133 (IAEDANS) in TnI,(3) Cys98 (IAEDANS) of TnC to Cys133 (IAE) of TnI,(4) Trp158 of TnI to Cys98 (IAEDANS) of TnC, and (6) Met25 (DNZ) of TnC to Cys133 (IAE) of TnI. Distance (1) in TnC was little affected when the isolated protein was complexed with TnI, whereas distance (2) in TnI increased by 6Å (29%) when TnI was incorporated into the binary complex. In the presence of EGTA, the six donor-acceptor separations (R) in the complex were in the range 28 to 57 Å based on κ 2= 2 3. Mg 2+ had only small effects on R, but Ca 2+ induced substantial increases or decreases of R in five of the six distances. These changes were not accompanied by significant changes in the axial depolarization of the fluorophores. The results indicate global structural perturbations of regions of the two proteins in the complex by Ca 2+ binding to the TnC, and suggest that large-scale movements of domains of troponin subunits may be the initial molecular events that occur in the transmission of the Ca 2+ signal in the regulation of contraction by calcium.