Free energy calculations on the stability of the 14-3-3ζ protein.
Free energy calculations on the stability of the 14-3-3ζ protein.
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DOI：
10.1016/j.bbapap.2017.11.012
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发表时间：
2018-03
期刊：
影响因子：
--
通讯作者：
Hritz J
中科院分区：
文献类型：
--
作者：
Jandova Z;Trosanova Z;Weisova V;Oostenbrink C;Hritz J
半胱氨酸的突变经常被引入，例如，在体外实验中避免形成非生理的分子间二硫键，或者在标记实验中保持特异性。当原来的半胱氨酸暴露在表面时，丙氨酸或丝氨酸通常是首选的，这通常不会改变蛋白质的整体稳定性。然而，选择蛋白质疏水核心半胱氨酸的最佳突变更具挑战性。在这项工作中，选择的14-3-3ζ的半胱氨酸突变体的稳定性进行了预测，并与实验测定的稳定性进行了比较。无论是计算预测还是实验验证点，都是突变C94A和C94S的显著不稳定。这种不稳定可以归因于疏水空腔的形成和亲水侧链的极性溶剂化。进一步研究了L12E、M78K双突变体的二聚化倾向。与天真的预期相反，这个双重突变体没有导致形成强大的盐桥，这是合理的，因为离子物种的优先溶剂化作用。再次，实验与计算一致，证实了双突变体的单聚化。总体而言，模拟数据与实验符合得很好，并为这一重要调控蛋白家族的稳定性和二聚化提供了额外的洞察力。
Mutations of cysteine are often introduced to e.g. avoid formation of non-physiological inter-molecular disulfide bridges in in-vitro experiments, or to maintain specificity in labeling experiments. Alanine or serine is typically preferred, which usually do not alter the overall protein stability, when the original cysteine was surface exposed. However, selecting the optimal mutation for cysteines in the hydrophobic core of the protein is more challenging. In this work, the stability of selected Cys mutants of 14-3-3ζ was predicted by free-energy calculations and the obtained data were compared with experimentally determined stabilities. Both the computational predictions as well as the experimental validation point at a significant destabilization of mutants C94A and C94S. This destabilization could be attributed to the formation of hydrophobic cavities and a polar solvation of a hydrophilic side chain. A L12E, M78K double mutant was further studied in terms of its reduced dimerization propensity. In contrast to naïve expectations, this double mutant did not lead to the formation of strong salt bridges, which was rationalized in terms of a preferred solvation of the ionic species. Again, experiments agreed with the calculations by confirming the monomerization of the double mutants. Overall, the simulation data is in good agreement with experiments and offers additional insight into the stability and dimerization of this important family of regulatory proteins.
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影响因子：
2.8
作者：
BEUTLER, TC;MARK, AE;VANGUNSTEREN, WF
通讯作者：
VANGUNSTEREN, WF
DOI：
10.1042/bj20091834
发表时间：
2010-03-15
期刊：
The Biochemical journal
影响因子：
--
作者：
Johnson C;Crowther S;Stafford MJ;Campbell DG;Toth R;MacKintosh C
通讯作者：
MacKintosh C
影响因子：
3.5
作者：
Nagano, N;Ota, M;Nishikawa, K
通讯作者：
Nishikawa, K
14-3-3-ζ 与整合素尾部相互作用的表征。
影响因子：
5.6
作者：
Bonet, Roman;Vakonakis, Ioannis;Campbell, Iain D.
通讯作者：
Campbell, Iain D.
影响因子：
2.9
作者：
BECKTEL, WJ;SCHELLMAN, JA
通讯作者：
SCHELLMAN, JA