Investigation of the acylation mechanism of class C beta-lactamase: pKa calculation, molecular dynamics simulation and quantum mechanical calculation |
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Authors: | Smriti Sharma Pradipta Bandyopadhyay |
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Institution: | (1) Centre for Computational Biology and Bioinformatics, School of Computational and Integrative Sciences, Jawaharlal Nehru University, New Delhi, 110067, India; |
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Abstract: | β-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. |
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