In-silico driven engineering of enantioselectivity of a penicillin G acylase towards active pharmaceutical ingredients

Grulich, Michal; Brezovský, Jan; Štěpánek, Václav; Palyzová, Andrea; Marešová, Helena; Zahradník, Jiří; Kyslíková, Eva; Kyslı́k, Pavel

doi:10.1016/j.molcatb.2016.11.014

Cited by 6 publications

(12 citation statements)

References 49 publications

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“…In this review we concentrated on the force field algorithm FoldX, which we have used by ourselves to create a more stable ω-transaminase [ 81 ]. The force field algorithm, which was originally created by Guerois et al became popular as webtool in 2005 by Schymkowitz et al and was refined to the currently last version FoldX 4.0 [ [82] , [83] , [84] ].…”

Section: Un/folding Energy Algorithmsmentioning

confidence: 99%

FoldX as Protein Engineering Tool: Better Than Random Based Approaches?

Buß

Rudat

Ochsenreither

2018

Computational and Structural Biotechnology Journal

194

125

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Improving protein stability is an important goal for basic research as well as for clinical and industrial applications but no commonly accepted and widely used strategy for efficient engineering is known. Beside random approaches like error prone PCR or physical techniques to stabilize proteins, e.g. by immobilization, in silico approaches are gaining more attention to apply target-oriented mutagenesis. In this review different algorithms for the prediction of beneficial mutation sites to enhance protein stability are summarized and the advantages and disadvantages of FoldX are highlighted. The question whether the prediction of mutation sites by the algorithm FoldX is more accurate than random based approaches is addressed.

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Section: Un/folding Energy Algorithmsmentioning

confidence: 99%

FoldX as Protein Engineering Tool: Better Than Random Based Approaches?

Buß

Rudat

Ochsenreither

2018

Computational and Structural Biotechnology Journal

194

125

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“…Conversely, the dynamics of the gating residues of the acyl-binding cavity of PGAs allow these enzymes to accommodate a much broader range of substrates including HSL, various amino acids, and penicillins. 51,54,85,87 Our QM/MM MD-based study revealed protein- and ligand-dependent differences in the dynamics of these gating residues, showing that complexes of aPGA with longer HSL substrates were more likely to sample closed conformations than the other PGA complexes included in the study. This observation agrees well with insights from previous studies suggesting a preference for closed states in the free enzyme and complexes with specifically recognized ligands, whereas open states are preferred for complexes with nonspecific ligands.…”

Section: Discussionmentioning

confidence: 81%

“…These studies have used both experimental techniques and a broad spectrum of molecular modeling methods including docking, MD simulations, and QM or QM/MM calculations. 53,57,69,70,85,95 Our study provides a complementary extension of the current knowledge of N-terminal serine hydrolases and particularly their QQ activity based on extensive QM/MM MD simulations. To our knowledge, comparable simulations have only been reported for cysteine N-terminal-hydrolases – closely related enzymes that behave differently to serine or threonine hydrolases because the catalytic cysteine preferentially adopts a zwitterionic form, 96,97 making an entirely analogous mechanism unlikely.…”

Section: Discussionmentioning

confidence: 87%

“…Mechanism-based geometric criteria defining the atomic arrangements required for the activity of N-terminal serine hydrolases can be expressed in terms of three criteria that must be satisfied simultaneously ( Figure 3 ): there must be a relatively short nucleophile attack distance (< 3.3 Å), it must be possible to form two stabilizing hydrogen bonds with oxyanion hole stabilizing residues, and the nucleophile attack angle must be ∼90°. 53,57,85…”

Section: Resultsmentioning

confidence: 99%

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Common Dynamic Determinants Govern Quorum Quenching Activity in N-terminal Serine Hydrolases

Surpeta

Grulich

Palyzová

et al. 2022

Preprint

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Due to the alarming global crisis of the growing microbial antibiotic resistance, investigation of alternative strategies to combat this issue has gained considerable momentum in the recent decade. A quorum quenching (QQ) process disrupts bacterial communication through so-called quorum sensing that enables bacteria to sense the cell density in the surrounding environment. Due to its indirect mode of action, QQ is believed to exert limited pressure on essential bacterial functions and consequently avoid inducing resistance. Although many enzymes are known to display the QQ activity towards various molecules used for bacterial signaling, the in-depth mechanism of their action is not well understood hampering their possible optimization for such exploitation. In this study, we compare the potential of three members of N-terminal serine hydrolases to degrade N-acyl homoserine lactones--signaling compounds employed by Gram-negative bacteria. Using molecular dynamics simulation of free enzymes and their complexes with two signaling molecules of different lengths, followed by quantum mechanics/molecular mechanics molecular dynamics simulation of their initial catalytic steps, we explored molecular details behind their QQ activities. We observed that all three enzymes were able to degrade bacterial signaling molecules following an analogous reaction mechanism. For the two investigated penicillin G acylases from Escherichia coli (ecPGA) and Achromobacter spp. (aPGA), we confirmed their putative activities experimentally hereby extending the set of known quorum quenching enzymes by these representatives of biotechnologically well-optimized enzymes. Interestingly, we detected enzyme- and substrate-depended differences among the three enzymes caused primarily by the distinct structure and dynamics of acyl-binding cavities. As a consequence, the first reaction step catalyzed by ecPGA with a longer substrate exhibited an elevated energy barrier due to a too shallow acyl-binding site incapable of accomodating this molecule in a required configuration. Conversely, unfavorable energetics on both reaction steps were observed for aPGA in complex with both substrates, conditioned primarily by the increased dynamics of the residues gating the entrance to the acyl-binding cavity. Finally, the energy barriers of the second reaction step catalyzed by Pseudomonas aeruginosa acyl-homoserine lactone acylase with both substrates were higher than in the other two enzymes due to distinct positioning of Arg297β. These discovered dynamic determinants constitute valuable guidance for further research towards designing robust QQ agents capable of selectively controlling the virulence of resistant bacteria species.

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“…Semisynthetic penicillins have been shown to have better properties than penicillin G and V, such as higher stability, easier absorption and lesser side effects and are also better candidates against the adaptive microbial resistance to antibiotics [42]. Large-scale production of semisynthetic antibiotics that are derived from penicillin are based on the condensation of the β-lactam nucleus with appropriate D-amino acid catalyzed by penicillin acylases (Scheme 1) [43][44][45][46].…”

Section: Synthesis Of Semisynthetic Penicillins By Penicillin Acylasesmentioning

confidence: 99%

Enzymes for pharmaceutical and therapeutic applications

Meghwanshi

Kaur

Verma

et al. 2020

Biotech and App Biochem

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Enzymes are highly efficient and selective biocatalysts, present in the living beings. They exist in enormous varieties in terms of the types of reactions catalyzed by them for instance oxidation–reduction, group transfers within the molecules or between the molecules, hydrolysis, isomerization, ligation, bond cleavage, and bond formation. Besides, enzyme based catalyses are performed with much higher fidelity, under mild reaction conditions and are highly efficient in terms of number of steps, giving them an edge over their chemical counter parts. The unique characteristics of enzymes makes them highly applicable fora number of chemical transformation reactions in pharmaceutical industries, such as group protection and deprotection, selective acylation and deacylation, selective hydrolysis, deracemization, kinetic resolution of racemic mixtures, esterification, transesterification, and many others. In this review, an overview of the enzymes, their production and their applications in pharmaceutical syntheses and enzyme therapies are presented with diagrams, reaction schemes and table for easy understanding of the readers.

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In-silico driven engineering of enantioselectivity of a penicillin G acylase towards active pharmaceutical ingredients

Cited by 6 publications

References 49 publications

FoldX as Protein Engineering Tool: Better Than Random Based Approaches?

FoldX as Protein Engineering Tool: Better Than Random Based Approaches?

Common Dynamic Determinants Govern Quorum Quenching Activity in N-terminal Serine Hydrolases

Enzymes for pharmaceutical and therapeutic applications

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