Analysis on the early events of protein folding at subzero temperatures

零下温度下蛋白质折叠的早期事件分析

• 批准号: 12680663
• 负责人: KIHARA Hiroshi
• 金额: $ 2.24万
• 依托单位: Kansai Medical University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12680663/
• 关键词: Protein folding; Compaction of protein; Formation rate of alpha-helix; Subzero temperature; Stopped-flow; x-ray solution scattering; フォールディング; ビータラクトグロブリン; アポミオグロビン; ユビキティン; アルフアービータ変換; 円偏光二色性

Protein takes (a) definite pathway(s) when it folds into its native form. It is so-called folding problem. One of the most important subject in the folding process is when and how the three major conformational formation takes place ; i.e. secondary structure formation, compaction and formation of the folding core.Protein folding starts at the time range of sub-micro-seconds. We reduced this rate to millisecond region, by decreasing temperature so as to slow down the folding process. With the use of stopped-flow technique combined with circular dichroism, fluorescence and X-ray scattering, we could monitor formation of alpha-helix, folding core formation and compaction. Results so far obtained are ; (1) formation rate of alpha-helix is too fast to be monitored even at -38C. (2) In case of bovine beta-lactogloblulin, we monitored a phase of alpha-helix increase at -28C. (3) In case of ubiquitin, the initial burs phase of alpha-helical formation was observed at -20C.From the findings above, it would be suggested that formation of alpha-helix is commonly observed at the early stage of the folding even in the case of beta-rich proteins.Results above are also very consistent with findings observed by temperature-jump method, and thus demonstrate the importance of our methodology ; cooling down the folding process.

当蛋白质折叠成天然形式时，蛋白质采用（a）确定的途径。这是所谓的折叠问题。折叠过程中最重要的主题之一是何时以及如何发生三个主要的构象形成。即折叠核心的二级结构形成，压实和形成。蛋白质折叠在亚微米秒的时间范围内开始。我们通过降低温度以减慢折叠过程来降低该速率。通过使用停止流动技术，结合了圆形二色性，荧光和X射线散射，我们可以监视α-螺旋形成，折叠核心形成和压实。到目前为止获得的结果是； （1）α-螺旋的形成速率太快了，即使在-38C下也无法监测。 （2）在牛β-乳腺癌蛋白的情况下，我们监测了-28C时α-螺旋增加的阶段。 （3）在泛素的情况下，在-20C处观察到α-螺旋形成的初始掌burs阶段。从上面的发现从上面的发现，建议的是，在折叠的早期阶段也观察到alpha-helix的形成，即使在beta蛋白富含蛋白质的情况下，也可以很好地观察到折叠的早期阶段。因此，在β-蛋白质中也非常一致。冷却折叠过程。

项目成果

Zhi-jie Qin, Dong-mei Hu, Lui Shimada, Tatsuo Nakagawa, Munehito Arai, Jun-Mei Zhou and Hiroshi Kihara.: "Refolding of β-Lactoglobulin Studied by Stopped-Flow Circular Dichroism at Subzero Temperatures"FEBS lett.. 507. 299-302 (2001)

秦志杰、胡冬梅、Lui Shimada、Tatsuo Nakakawa、Munehito Arai、Jun-Mei Zhou 和 Hiroshi Kihara.：“在零下温度下通过停流圆二色性研究 β-乳球蛋白的重折叠”FEBS lett.. 507 .299-302 (2001)

Qin et al.: "Refolding of β-lactoglobulin studied by stopped-flow ciecular dichroism at subzero temperatures"FEBS Lett.. 507. 299-302 (2001)

秦等人：“在零度以下温度下通过停流环二色性研究β-乳球蛋白的重折叠”FEBS Lett.. 507. 299-302 (2001)

Oin et al.: "Refolding of β-lactoglobulin studied by stopped-flow ciecular dichroism at subzero temperatures"FEBS Lett.. 507. 299-302 (2001)

Oin 等人：“在零下温度下通过停流环二色性研究 β-乳球蛋白的重折叠” FEBS Lett.. 507. 299-302 (2001)

A.A.Timchenko et al.: "GroES co-chaperonin small-angle X-ray scattering study shows ring orifice increase in solution"FEBS Lett.. 471. 211-214 (2000)

A.A.Timchenko 等人：“GroES 辅助伴侣小角 X 射线散射研究显示溶液中环形孔口增加”FEBS Lett.. 471. 211-214 (2000)