Local mobility within villin 14T probed via heteronuclear relaxation measurements and a reduced spectral density mapping.
Local mobility within villin 14T probed via heteronuclear relaxation measurements and a reduced spectral density mapping.
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通过异核弛豫测量和降低的谱密度映射探测 villin 14T 内的局部迁移率。
DOI：
10.1021/bi951933o
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Wagner,G
中科院分区：
文献类型：
--
作者：
Markus,MA;Dayie,KT;Matsudaira,P;Wagner,G
绒毛蛋白14 T是肌动蛋白切割和捆绑蛋白绒毛蛋白的代表性结构域，结合钙离子和肌动蛋白单体。为了开始理解迁移率对绒毛-钙和绒毛-肌动蛋白相互作用的贡献，测量了溶液中15 N标记的绒毛14 T的酰胺氮和质子的磁化弛豫速率。虽然我们已经测量了完整的频谱密度图所需的一套完整的速率，在准确测量的反相相干和两个自旋顺序的弛豫率的困难，使我们考虑减少映射形式主义。从简化的谱密度图，无模型分析，或直接从测量的Nx，y弛豫速率，局部变化的流动性沿着骨架的绒毛14 T已被揭示。快速运动不仅在氨基和羧基末端很明显，而且在中心β折叠的β4和β5链之间的转角以及螺旋α3和β7链之间的转角也很明显。对于股β2和股β3之间的转弯，建议较慢的运动。微秒到毫秒的时间尺度上的运动已被直接探测通过检查质子横向弛豫速率的依赖性的自旋锁定场强度。Leu 11对电场强度有很强的依赖性，表明构象交换的时间常数为125 ± 69 μs。在肌动蛋白结合界面的骨干似乎是相当刚性。
Villin 14T, a representative domain from the actin severing and bundling protein villin, binds calcium ions and actin monomers. To begin to understand the contributions of mobility to the villin−calcium and villin−actin interactions, relaxation rates for magnetization involving the amide nitrogens and protons have been measured for15N-labeled villin 14T in solution. Although we have measured the complete set of rates required for a full spectral density map, difficulties in the accurate measurement of relaxation rates for antiphase coherence and two-spin order led us to consider a reduced mapping formalism. From the reduced spectral density map, a model-free analysis, or directly from the measured Nx,yrelaxation rates, local variations in mobility along the backbone of villin 14T have been revealed. Fast motions are evident not only at the amino and carboxyl termini but also in the turn between strands β4 and β5 of the central β-sheet and in the turn between helix α3 and strand β7. Slower motions are suggested for the turn between strands β2 and β3. Motions on the microsecond to millisecond time scale have been probed directly by examining the dependence of the proton transverse relaxation rate on the spin-locking field strength. Leu11shows a strong dependence on field strength, implying conformational exchange with a time constant of 125 ± 69 μs. The backbone at the actin-binding interface appears to be rather rigid.