Thermodynamics of a protein acylation: activation of Escherichia coli hemolysin toxin.
Thermodynamics of a protein acylation: activation of Escherichia coli hemolysin toxin.
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蛋白质酰化的热力学：大肠杆菌溶血素毒素的激活。
DOI：
10.1021/bi048479l
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发表时间：
2005
期刊：
影响因子：
--
通讯作者：
Ernst-Fonberg,MLou
中科院分区：
文献类型：
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作者：
Worsham,LesaMS;Langston,KeishaG;Ernst-Fonberg,MLou
HlyC，溶血素活化赖氨酸酰基转移酶，催化大肠杆菌溶血素原（prohemolysin，proHlyA）在1024个氨基酸一级结构的第564和690个赖氨酸残基的ε-氨基上的酰化（来自酰基-酰基载体蛋白[ACP]），形成溶血素（hemolysin，HlyA）。等温滴定量热法用于测量proHlyA衍生结构的蛋白质酰化的热力学性质，通过大量缺失和酰化位点分离成两种不同的肽和酰化位点的定点突变分析来改变。由HlyC催化的proHlyA衍生的蛋白质的酰化总体上是由负焓驱动的放热反应。该反应由两个部分反应组成，其动力学符合乒乓机理。第一种是酰基-HlyC中间体的形成，它是熵驱动的，最有可能是由酰基-ACP和HlyC之间的非共价复合物形成;随后是酰基-ACP驱动的酰基转移，导致酰基-HlyC和ACPSH产物形成。第二部分反应是从酰基酶中间体到最终酰基受体（proHlyA衍生物）的能量上不利的酰基转移。总的来说，由HlyC催化的proHlyA衍生蛋白的酰化由酰基酶中间体反应的能量学驱动。在两个酰化位点中，与proHlyA K564相当的位点的完整性对酰化反应热力学稳定性更重要。
HlyC, hemolysin-activating lysine-acyltransferase, catalyses the acylation (from acyl−acyl carrier protein [ACP]) ofEscherichia coliprohemolysin (proHlyA) on the ε-amino groups of specific lysine residues, 564 and 690 of the 1024 amino acid primary structure, to form hemolysin (HlyA). Isothermal titration calorimetry was used to measure the thermodynamic properties of the protein acylation of proHlyA-derived structures, altered by substantial deletions and separation of the acylation sites into two different peptides and site directed mutation analyses of acylation sites. Acylation of proHlyA-derived proteins catalyzed by HlyC was overall an exothermic reaction driven by a negative enthalpy. The reaction, whose kinetics are compatible to a ping-pong mechanism, is composed of two partial reactions. The first, the formation of an acyl−HlyC intermediate, was entropically driven, most likely by noncovalent complex formation between acyl-ACP and HlyC; enthalpy-driven acyl transfer followed, resulting in acyl−HlyC and ACPSH product formation. The second partial reaction was an energetically unfavorable acyl transfer from acyl-enzyme intermediate to the final acyl acceptor, a proHlyA derivative. Overall the acylation of proHlyA-derived proteins catalyzed by HlyC was driven by the energetics of the acyl enzyme intermediate reaction. Of the two acylation sites, intactness of the site equivalent to proHlyA K564 was more important for acylation reaction thermodynamic stability.