Reversible and fast association equilibria of a molecular chaperone, gp57A, of bacteriophage T4
Reversible and fast association equilibria of a molecular chaperone, gp57A, of bacteriophage T4
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DOI：
10.1016/s0006-3495(03)74683-9
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发表时间：
2003-10-01
影响因子：
3.4
通讯作者：
Arisaka, F
中科院分区：
文献类型：
--
作者：
Ali, SA;Iwabuchi, N;Arisaka, F
采用分析超离心、差示扫描微热法和停流圆二色性（CD）等方法研究了T4噬菌体中促进长、短尾纤维形成的分子伴侣蛋白gp57A的结合力，建立了该蛋白的结合力方案。Gp57A是一种低聚α -螺旋蛋白，含有79个氨基酸。通过Lamm方程解的直接边界建模以及结合关联方案的更详细的边界分析，我们得出结论，至少有三种gp57A的寡聚物种处于可逆和快速的关联平衡状态，并且3(mer)(mer)(mer)(mer)(-6)（-12）模型最能描述数据。另一方面，差示扫描微量热法显示了高度可逆的解离/变性两步转变，两者都伴随着CD在222 nm处的下降。熔点曲线分析表明，熔点曲线符合6(mer)(mer)(mer)(mer)(-3)（-1）模型。停流CD测量的gp57A的再折叠/结合动力学与从三聚体到六聚体的双分子反应之前在死区快速形成α -螺旋的解释一致。三聚体或六聚体可能是gp57A的功能性低聚体状态。
The association of a molecular chaperone, gp57A, of bacteriophage T4, which facilitates formation of the long and short tail fibers, was investigated by analytical ultracentrifugation, differential scanning microcalorimetry, and stopped-flow circular dichroism (CD) to establish the association scheme of the protein. Gp57A is an oligomeric alpha-helix protein with 79 amino acids. Analysis of the sedimentation velocity data by direct boundary modeling with Lamm equation solutions together with a more detailed boundary analysis incorporating association schemes led us to conclude that at least three oligomeric species of gp57A are in reversible and fast association equilibria and that a 3(mer)(mer)(mer)(-6)(-12) model described the data best. On the other hand, differential scanning microcalorimetry revealed a highly reversible two-step transition of dissociation/denaturation, both of which accompanied decrease in CD at 222 nm. The melting curve analysis revealed that it is consistent with a 6(mer)(mer)(mer)(-3)(-1) model. The refolding/association kinetics of gp57A measured by stopped-flow CD was consistent with the interpretation that the bimolecular reaction from trimer to hexamer was preceded by a fast alpha-helix formation in the dead-time. Trimer or hexamer is likely the functional oligomeric state of gp57A.