Solid-state NMR studies on the supersecondary structures in fibrous proteins

纤维蛋白超二级结构的固态核磁共振研究

• 批准号: 07651103
• 负责人: YOSHIMIZU Hiroaki
• 金额: $ 1.6万
• 依托单位: Nagoya Institute of Technology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07651103/
• 关键词: fibrous proteins; supeseconday structures; coild-coil structure; alpha-helix; conformation; soild-state NMR; chemical shift; wool keratin

^<13>C CP/MAS NMR spectra of tropomyosin were measured in order to elucidate the higher order structure through the ^<13>C NMR chemical shifts of the amino acid residues and their mobility. The higher order structure of tropomyosin is a coiled-coil structure, and contains two different sites which are characterized as the internal and external sites. The ^<13>C signal which appear at 15-17 ppm can be easily assigned to the Ala C^<beta> carbons, and it can be seen that this signal was consisted of two peaks. The peak at 15.8 ppm have a longer T_1 value than those at 16.7 ppm. This is indicated that the former carbons are more mobile than the latter carbons. Therefore, the two peaks at 15.8 and 16.7 ppm are assigned to the Ala C^<beta> carbons in the extermal and internal sites of the coiled-coil structure, respectively. Wool keratin can be divided into three main fractions after reducing disulfide bonds and protection of the resulting thiol groups with iodoacetic acid to form S-carboxymethyl kerateine (SCMK). From the ^<13>C CP/MAS NMR spectra of wool and SCMK,it was suggested that the coiled-coil structure exists because the ^<13>C chemical shift values of Ala C^<beta> carbons in these samples coincide in experimental error with that of the Ala residue located in the internal site of the coiled-coil structure in tropomyosin. The conformational analysis using the conformation-dependent ^<13>C CP/MAS NMR chemical shifts were applied to estimate the conformational transition of SCMK by stretching, heating, or steam-treating. The beta-sheet form appears by stretching and steam-treating. For the heated SCMK,it can be said that the random coil form appears. However, the fact that the a-helix form remains to an appreciable extent in the heated one although the random coil form appears, suggests that only the packing of the coiled-coil structure in the SCMK is disrupted by heating, but secondary structure being retained.

测量原肌球蛋白的 13 C CP/MAS NMR谱，以便通过氨基酸残基的 13 C NMR化学位移及其迁移率阐明高阶结构。原肌球蛋白的高级结构是卷曲螺旋结构，并且包含两个不同的位点，其特征为内部位点和外部位点。在15-17ppm处出现的^ 13 C信号可以容易地归属于Ala C ^ β 碳，并且可以看出该信号由两个峰组成。 15.8 ppm 处的峰比 16.7 ppm 处的峰具有更长的 T_1 值。这表明前一种碳比后一种碳更易移动。因此，15.8和16.7ppm处的两个峰分别归属于卷曲螺旋结构的外部和内部位点中的Ala C 16 碳。羊毛角蛋白在还原二硫键并用碘乙酸保护所得硫醇基团后可分为三个主要部分，形成S-羧甲基角蛋白（SCMK）。从羊毛和SCMK的^<13>C CP/MAS NMR谱来看，由于这些样品中Ala C^<β碳的^<13>C化学位移值一致，表明存在卷曲螺旋结构。与位于原肌球蛋白卷曲螺旋结构内部位点的丙氨酸残基的实验误差。应用使用构象依赖性13 C CP/MAS NMR化学位移的构象分析来估计SCMK通过拉伸、加热或蒸汽处理的构象转变。通过拉伸和蒸汽处理形成β-折叠形式。对于加热的SCMK，可以说出现了无规线圈形式。然而，尽管出现了无规卷曲形式，但加热后的α螺旋形式仍保留在相当大的程度上，这一事实表明，只有SCMK中卷曲卷曲结构的堆积因加热而被破坏，但二级结构被保留。

项目成果

高分子学会編: "高分子実験学5 高分子の構造(1)磁気共鳴法" 共立出版株式会社, 572 (1995)

高分子学会编：《高分子实验5：高分子的结构(1)磁共振法》共立出版572(1995)

Hiroaki Yoshimizu: Section 4.5.1 in "The Structures of Polymers (1) Hagnetic Resonance Spectroscopy". Edited by the Sociery of Polymer Science, Japan, Kyoritsu Shuppan Co.Ltd., (1995)

Hiroaki Yoshimizu：《聚合物的结构（1）共振光谱》第4.5.1节。