Method-Unifying View of Loop-Formation Kinetics in Peptide and Protein Folding.
Method-Unifying View of Loop-Formation Kinetics in Peptide and Protein Folding.
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肽和蛋白质折叠中环形成动力学的方法统一观点
DOI：
10.1021/acs.jpcb.8b00879
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发表时间：
2018
期刊：
影响因子：
--
通讯作者：
Schwarzlose
中科院分区：
文献类型：
--
作者：
D’Souza;Schwarzlose
蛋白质折叠可以被描述为事件的概率性连续，其中肽链形成由特定氨基酸残基接触闭合的环，本文称为环节点。为了测量环速率，已经引入了几种光物理方法，其中在选定的链位置处并入一对光学活性探针，并且激发的探针在与第二探针碰撞时经历接触淬灭（CQ）。猝灭机制涉及三重态-三重态能量转移、光诱导电子转移和碰撞诱导荧光猝灭，其中Dbo（与2，3-二氮杂双环[2.2.2]辛烷缀合的天冬酰胺残基）的荧光被色氨酸猝灭。然而，这三种CQ技术提供的环路速率之间的差异仍然没有得到解决。在分析这种差异，我们现在报告两个短距离FRET方法，其中Dbo作为能量受体与色氨酸和naphthylalanine，两个供体有很大不同的荧光寿命分别为1.3和33 ns的组合。尽管不同的淬灭机制，从FRET和CQ方法的速率，令人惊讶的是，相当的幅度。FRET和CQ数据的这种组合导致了一个统一的物理模型，并得出结论，即折叠反应中的环形成速率不仅随构成链的残基的种类和数量而变化，而且特别是随构成环节点的残基的大小和性质而变化。
Protein folding can be described as a probabilistic succession of events in which the peptide chain forms loops closed by specific amino acid residue contacts, herein referred to as loop nodes. To measure loop rates, several photophysical methods have been introduced where a pair of optically active probes is incorporated at selected chain positions and the excited probe undergoes contact quenching (CQ) upon collision with the second probe. The quenching mechanisms involved triplet–triplet energy transfer, photoinduced electron transfer, and collision-induced fluorescence quenching, where the fluorescence of Dbo, an asparagine residue conjugated to 2,3-diazabicyclo[2.2.2]octane, is quenched by tryptophan. The discrepancy between the loop rates afforded from these three CQ techniques has, however, remained unresolved. In analyzing this discrepancy, we now report two short-distance FRET methods where Dbo acts as an energy acceptor in combination with tryptophan and naphtylalanine, two donors with largely different fluorescence lifetimes of 1.3 and 33 ns, respectively. Despite the different quenching mechanisms, the rates from FRET and CQ methods were, surprisingly, of comparable magnitude. This combination of FRET and CQ data led to a unifying physical model and to the conclusion that the rate of loop formation in folding reactions varies not only with the kind and number of residues that constitute the chain but also in particular with the size and properties of the residues that constitute the loop node.