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首页> 外文学位 >Nuclear magnetic resonsance studies of side-chain motions in calbindin D(9k): The role of conformational dynamics in protein stability and calcium binding.
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Nuclear magnetic resonsance studies of side-chain motions in calbindin D(9k): The role of conformational dynamics in protein stability and calcium binding.

机译：钙结合蛋白D（9k）中侧链运动的核磁共振研究：构象动力学在蛋白质稳定性和钙结合中的作用。

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摘要

An accurate understanding of the role of conformational dynamics in proteins requires data at multiple timescales and sites within the protein of interest. Considerable progress has been achieved in characterizing the picosecond-to-nanosecond (ps-ns) dynamics of the protein backbone via NMR relaxation measurements of the 15N nucleus. More recent developments in the measurement of 2H quadrupolar relaxation rates are enabling an extensive characterization of the dynamics in methyl-containing side-chains as well. The aim of the present study is to characterize the effects of Ca2+ binding on the side-chain dynamics of the protein calbindin D 9k. Calbindin is a small (∼8.7 kD), single domain protein of the EF-hand family. It contains two Ca2+ binding sites that exhibit high positive cooperativity. Longitudinal, transverse, quadrupolar order, transverse antiphase and double quantum relaxation rates are reported for both the apo (Ca2+-free) and Ca2+-loaded states of the protein at two magnetic field strengths. The relatively large size of the data set allows for a detailed analysis of the underlying conformational dynamics by spectral density mapping and model-free fitting procedures. The results indicate that a methyl group's distance from the Ca2+ binding sites is a significant determinant of its conformational dynamics. Several methyl groups segregate into two limiting classes, one proximal and the other distal to the binding sites. Methyl groups in these two classes respond differently to Ca2+ binding, both in terms of the timescale and amplitude of their fluctuations. Ca2+ binding elicits a partial immobilization among methyl groups in the proximal class, which is consistent with previous studies of calbindin's backbone dynamics. The distal class, however, exhibits a trend that could not be inferred from the backbone data in that its mobility actually increases with Ca2+ binding. We have introduced the term polar dynamics to describe this type of organization across the molecule. The trend may represent an important mechanism by which calbindin achieves high affinity binding while minimizing the corresponding conformational entropy loss.
机译：对构象动力学在蛋白质中的作用的准确理解需要在目标蛋白质内多个时间尺度和位点的数据。通过15N核的NMR弛豫测量，在表征蛋白主链的皮秒至纳秒（ps-ns）动力学特性方面已经取得了显着进展。测量2H四极弛豫速率的最新进展也使得能够广泛表征含甲基侧链的动力学。本研究的目的是表征Ca 2+结合对钙结合蛋白D 9k的侧链动力学的影响。钙结合蛋白是EF手家族的小（约8.7kD）单结构域蛋白。它包含两个具有高正协同性的Ca2 +结合位点。据报道，在两个磁场强度下，蛋白质的载脂蛋白（无Ca2 +）和Ca2 +加载态的纵向，横向，四极顺序，横向反相和双量子弛豫速率都得到了报道。数据集的相对较大的大小允许通过光谱密度映射和无模型拟合程序对潜在的构象动力学进行详细分析。结果表明，甲基与Ca 2+结合位点的距离是其构象动力学的重要决定因素。几个甲基分为两个限制类别，一个位于结合位点的近端，另一个位于结合位点的远端。就时间尺度和波动幅度而言，这两类甲基对Ca 2+结合的反应不同。 Ca 2+结合引起近端类别的甲基之间的部分固定，这与钙调蛋白骨架动力学的先前研究一致。然而，远端类表现出无法从骨干数据推断出的趋势，因为其活动性实际上随Ca2 +结合而增加。我们引入了极地动力学一词来描述分子中这种类型的组织。该趋势可能代表一种重要的机制，通过该机制，钙结合蛋白可实现高亲和力结合，同时最大程度地减少相应的构象熵损失。

著录项

  • 作者

    Johnson, Eric.;

  • 作者单位

    University of Cincinnati.;

  • 授予单位 University of Cincinnati.;
  • 学科 Chemistry Biochemistry.; Biophysics General.
  • 学位 Ph.D.
  • 年度 2005
  • 页码 147 p.
  • 总页数 147
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 生物化学;生物物理学;
  • 关键词

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