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Rationalizing perhydrolase activity of aryl-esterase and subtilisin Carlsberg mutants by molecular dynamics simulations of the second tetrahedral intermediate state

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Abstract

The perhydrolysis reaction in hydrolases is an important example of catalytic promiscuity and has many potential industrial applications. The mechanisms of perhydrolase activity of a subtilisin Carlsberg mutant and of an aryl-esterase mutant have been investigated using classical molecular dynamics simulations of the second tetrahedral intermediate (TI) state. The simulations demonstrated that hydrogen bonding between the second TI of the perhydrolysis reaction is possible in the mutants but not wild type. The stabilization by hydrogen bonds was specific for the perhydrolysis intermediate and either no hydrogen bonding or only weakened hydrogen bonding to the second TI state of the hydrolysis reaction was observed. Furthermore, a significant hindrance to the formation of the catalytically important hydrogen bond between His64 and Ser221 in the catalytic triad by competing hydrogen bonds was found for the subtilisin mutant but not wild type enzyme in case of the hydrolysis intermediate. The opposite was observed in case of the perhydrolysis intermediate. The result offers a qualitative explanation for the overall reduced hydrolysis activity of the subtilisin mutant. In addition, the simulations also explain qualitatively the perhydrolysis activity of the enzyme variants and may be helpful for designing enzyme mutants with further improved perhydrolysis activity.

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Acknowledgments

This work was performed using the computational resources of the CLAMV (Computer Laboratories for Animation, Modeling and Visualization) at Jacobs University Bremen and supercomputer resources of the EMSL (Environmental Molecular Science Laboratories) at the PNNL (Pacific Northwest National Laboratories).

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Author notes

  1. Martin Zacharias

    Present address: Physics Department, Technical University Munich, James Franck Str. 1, 85747, Garching, Germany

Authors and Affiliations

  1. School of Engineering and Science, Jacobs University, Campus Ring 1, 28759, Bremen, Germany

    Ljubica Vojcic, Dragana Despotovic, Radivoje Prodanovic & Martin Zacharias

  2. Institute of Organic Chemistry, University of Würzburg, 97074, Am Hubland, Würzburg

    Wook Lee

  3. WRC-Biotechnology, Henkel AG & Co. KGaA, 40191, Düsseldorf, Germany

    Karl-Heinz Maurer

  4. RWTH Aachen, Worringerweg 1, 52074, Aachen, Germany

    Ulrich Schwaneberg

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  1. Wook Lee
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  2. Ljubica Vojcic
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Correspondence to Martin Zacharias.

Additional information

Dedicated to Professor Sandor Suhai on the occasion of his 65th birthday and published as part of the Suhai Festschrift Issue.

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Lee, W., Vojcic, L., Despotovic, D. et al. Rationalizing perhydrolase activity of aryl-esterase and subtilisin Carlsberg mutants by molecular dynamics simulations of the second tetrahedral intermediate state. Theor Chem Acc 125, 375–386 (2010). https://doi.org/10.1007/s00214-009-0611-3

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