Investigation of the acylation mechanism of class C beta-lactamase: pKa calculation, molecular dynamics simulation and quantum mechanical calculation
Investigation of the acylation mechanism of class C beta-lactamase: pKa calculation, molecular dynamics simulation and quantum mechanical calculation
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C类β-内酰胺酶酰化机制研究：pKa计算、分子动力学模拟和量子力学计算
DOI：
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发表时间：
2012
影响因子：
2.2
通讯作者：
P. Bandyopadhyay
中科院分区：
文献类型：
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作者：
Smriti Sharma;P. Bandyopadhyay
摘要β-内酰胺酶是细菌对β-内酰胺类抗生素具有防御作用的酶。β-内酰胺酶通过涉及酰化和脱酰化步骤的两步机制切割抗生素的β-内酰胺环。虽然C类β-内酰胺酶已被广泛研究，但其作用机制的细节在分子水平上还不清楚。本研究采用pKa计算、分子动力学（MD）模拟和量子力学（QM）计算对C类β-内酰胺酶的酰化反应机理进行了研究。丝氨酸64（Ser 64）是攻击β-内酰胺环的活性位点残基。在这项研究中，我们考虑了亲核试剂Ser 64激活的三种可能情况，其中激活碱基是（1）酪氨酸150（Tyr 150），（2）赖氨酸67（Lys 67）或（3）底物。从pKa计算中，我们发现Tyr 150和Lys 67可能在酶和底物之间的前共价复合物中保持其质子化状态，尽管它们作为激活剂的作用需要它们处于去质子化状态。发现对于大部分模拟，底物的羧酸根基团保持接近Ser 64。采用酶活性中心的大截短模型计算了底物从丝氨酸64上夺氢的能垒，发现与实验能垒接近，表明底物可以启动C类β-内酰胺酶的酰化反应.
Abstractβ-Lactamases are bacterial enzymes that act as a bacterial defense system against β-lactam antibiotics. β-Lactamase cleaves the β-lactam ring of the antibiotic by a two step mechanism involving acylation and deacylation steps. Although class C β-lactamases have been investigated extensively, the details of their mechanism of action are not well understood at the molecular level. In this study, we investigated the mechanism of the acylation step of class C β-lactamase using pKa calculations, molecular dynamics (MD) simulations and quantum mechanical (QM) calculations. Serine64 (Ser64) is an active site residue that attacks the β-lactam ring. In this study, we considered three possible scenarios for activation of the nucleophile Ser64, where the activation base is (1) Tyrosine150 (Tyr150), (2) Lysine67 (Lys67), or (3) substrate. From the pKa calculation, we found that Tyr150 and Lys67 are likely to remain in their protonated states in the pre-covalent complex between the enzyme and substrate, although their role as activator would require them to be in the deprotonated state. It was found that the carboxylate group of the substrate remained close to Ser64 for most of the simulation. The energy barrier for hydrogen abstraction from Ser64 by the substrate was calculated quantum mechanically using a large truncated model of the enzyme active site and found to be close to the experimental energy barrier, which suggests that the substrate can initiate the acylation mechanism in class C β-lactamase.
影响因子：
4.1
作者：
Majiduddin, FK;Materon, IC;Palzkill, TG
通讯作者：
Palzkill, TG
影响因子：
7.3
作者：
Powers, RA;Shoichet, BK
通讯作者：
Shoichet, BK
影响因子：
2.9
作者：
Wang, Junmei;Wang, Wei;Case, David A.
通讯作者：
Case, David A.