The gramicidin dimer shows both EX1 and EX2 mechanisms of H/D exchange.
The gramicidin dimer shows both EX1 and EX2 mechanisms of H/D exchange.
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DOI：
10.1016/j.jasms.2009.05.017
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发表时间：
2009-10
影响因子：
3.2
通讯作者：
Gross ML
中科院分区：
文献类型：
--
作者：
Chitta RK;Rempel DL;Gross ML
我们描述了使用 H/D 酰胺交换和电喷雾电离质谱法来研究有机溶剂中的十五肽短杆菌肽作为蛋白质自缔合的模型。在甲醇-OD 中，肽中的所有活性 H 在 5 分钟内交换为 D，表明单体/二聚体平衡向快速交换单体转移。然而，正丙醇-OD 中的 H/D 交换显示出部分受保护的短杆菌肽，该短杆菌肽缓慢转化为交换几乎所有活性氢的第二种物质，表明 H/D 交换的 EX1 动力学。我们认为这种行为是由于正丙醇中的展开速率比甲醇中的展开速率慢的结果。二聚体展开的速率常数是部分受保护物种的消失速率，它与文献报道的值在两倍以内一致。二聚体重折叠的速率常数可以根据解折叠速率常数与二聚体亲和力常数的比率来确定，这是我们在早期研究中确定的。展开活化能为 20 kcal mol-1，通过作为温度函数的交换实验确定。为了在比正丙醇更疏水的介质中研究短杆菌肽，我们测量了其在磷脂囊泡中的 H/D 交换动力学，并发现了不同的 H/D 酰胺交换行为。短杆菌肽是一种不寻常的肽二聚体，根据溶剂的不同，其 H/D 交换可以同时表现出 EX1 和 EX2 机制。
We describe the use of H/D amide exchange and electrospray ionization mass spectrometry to study, in organic solvents, the pentadecapeptide gramicidin as a model for protein self association. In methanol-OD, all active H’s in the peptide exchange for D within 5 min, indicating a monomer/dimer equilibrium that is shifted towards the fast-exchanging monomer. H/D exchange in n-propanol-OD, however, showed a partially protected gramicidin that slowly converts to a second species that exchanges nearly all the active hydrogens, indicating EX1 kinetics for the H/D exchange. We propose that this behavior is the result of slower rate of unfolding in n-propanol compared to that in methanol. The rate constant for the unfolding of the dimer is the rate of disappearance of the partially protected species, and it agrees within a factor of two with a value reported in literature. The rate constant of dimer refolding can be determined from the ratio of the rate constant for unfolding and the affinity constant for the dimer, which we determined in an earlier study. The unfolding activation energy is 20 kcal mol-1, determined by performing the exchange experiments as a function of temperature. To study gramicidin in an even more hydrophobic medium than n-propanol, we measured its H/D exchange kinetics in a phospholipids vesicle and found a different H/D amide exchange behavior. Gramicidin is an unusual peptide dimer that can exhibit both EX1 and EX2 mechanisms for its H/D exchange, depending on the solvent.
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DOI：
10.1073/pnas.83.17.6258
发表时间：
1986-09-01
影响因子：
11.1
作者：
GRASBERGER, B;MINTON, AP;METZGER, H
通讯作者：
METZGER, H
影响因子：
1.8
作者：
BARONE, G;GIANCOLA, C
通讯作者：
GIANCOLA, C
影响因子：
7.2
作者：
Liu, D;Huang, Z
通讯作者：
Huang, Z
影响因子：
56.9
作者：
MIRANKER, A;ROBINSON, CV;DOBSON, CM
通讯作者：
DOBSON, CM
影响因子：
15
作者：
SARGES, R;WITKOP, B
通讯作者：
WITKOP, B