PROLINE ISOMERISM LEADS TO MULTIPLE FOLDED CONFORMATIONS OF CALBINDIN D9K - DIRECT EVIDENCE FROM TWO-DIMENSIONAL H-1-NMR SPECTROSCOPY
PROLINE ISOMERISM LEADS TO MULTIPLE FOLDED CONFORMATIONS OF CALBINDIN D9K - DIRECT EVIDENCE FROM TWO-DIMENSIONAL H-1-NMR SPECTROSCOPY
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DOI：
10.1073/pnas.86.7.2195
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发表时间：
1989-04-01
影响因子：
11.1
通讯作者：
FORSEN, S
中科院分区：
文献类型：
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作者：
CHAZIN, WJ;KORDEL, J;FORSEN, S
通过二维1H核磁共振光谱对钙结合蛋白D9 k进行了完整的分析，确定了溶液中折叠蛋白质存在两种构象。在整个1H NMR谱中观察到3：1比率的良好分辨的主共振和次共振。二维交换实验表明，主要和次要物种的平衡过程有关。短质子-质子距离沿着的肽骨架，确定了二维核Overhauser效应光谱分析，提供了明确的证据表明，这两种形式的折叠蛋白质的不同之处仅在于异构化状态的Gly-42和Pro-43之间的肽键。因此，Pro-43的顺反异构被直接鉴定为溶液中折叠蛋白质的多种构象的原因。此外，当Pro-43突变为甘氨酸残基时，没有多种构象的迹象。这些结果提供了证据的可能性的构象异质性的天然状态的球状蛋白质。
A complete analysis of calbindin D9k by two-dimensional 1H nuclear magnetic resonance spectroscopy has established the existence of two conformations for the folded protein in solution. Well-resolved major and minor resonances in a ratio of 3:1 are observed throughout the 1H NMR spectrum. Two-dimensional exchange experiments show that the major and minor species are related by an equilibrium process. Analysis of short proton-proton distances along the peptide backbone, identified by two-dimensional nuclear Overhauser effect spectroscopy, provides unambiguous evidence that the two forms of the folded protein differ only in the isomerization state of the peptide bond between Gly-42 and Pro-43. Cis-trans isomerism of Pro-43 is thereby directly identified as the cause of multiple conformations for the folded protein in solution. In addition, when Pro-43 is mutated to a glycine residue there is no indication of multiple conformations. These results provide evidence for the possiblity of conformational heterogeneity in the native state of globular proteins.