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Computational study of elements of stability of a four-helix bundle protein biosurfactant

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Abstract

Biosurfactants are surface-active molecules produced principally by microorganisms. They are a sustainable alternative to chemically-synthesized surfactants, having the advantages of being non-toxic, highly functional, eco-friendly and biodegradable. However they are currently only used in a few industrial products due to costs associated with production and purification, which exceed those for commodity chemical surfactants. DAMP4, a member of a four-helix bundle biosurfactant protein family, can be produced in soluble form and at high yield in Escherichia coli, and can be recovered using a facile thermal phase-separation approach. As such, it encompasses an interesting synergy of biomolecular and chemical engineering with prospects for low-cost production even for industrial sectors. DAMP4 is highly functional, and due to its extraordinary thermal stability it can be purified in a simple two-step process, in which the combination of high temperature and salt leads to denaturation of all contaminants, whereas DAMP4 stays stable in solution and can be recovered by filtration. This study aimed to characterize and understand the fundamental drivers of DAMP4 stability to guide further process and surfactant design studies. The complementary use of experiments and molecular dynamics simulation revealed a broad pH and temperature tolerance for DAMP4, with a melting point of 122.4 °C, suggesting the hydrophobic core as the major contributor to thermal stability. Simulation of systematically created in silico variants of DAMP4 showed an influence of number and location of hydrophilic mutations in the hydrophobic core on stability, demonstrating a tolerance of up to three mutations before a strong loss in stability occurred. The results suggest a consideration of a balance of stability, functionality and kinetics for new designs according to their application, aiming for maximal functionality but at adequate stability to allow for cost-efficient production using thermal phase separation approaches.

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Acknowledgments

The authors would like to gratefully acknowledge funding by an Australian Research Council Discovery Grant (DP120103683). A.S. was financially supported by an UQI Tuition Fee Scholarship and an AIBN RHD Living Scholarship. A.P.J.M. thanks the Queensland Government award of the 2010 Smart State Premier’s Fellowship. We acknowledge Lei Yu for executing DAMP4 expression and purification. Further, we would like to thank the HPC support from the University of Queensland Research Computing Centre (RCC), the Queensland Cyber Infrastructure Foundation (QCIF) and NCI National Facility for their support and the supercomputer resource allocation.

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Authors and Affiliations

  1. Centre for Biomolecular Engineering, Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, St Lucia, QLD, 4072, Australia

    Andrea Schaller, Natalie K. Connors, Mirjana Dimitrijev Dwyer & Anton P. J. Middelberg

  2. Section IV: Biomolecular Separation Engineering, Institute of Process Engineering in Life Sciences, Karlsruhe Institute of Technology, 76131, Karlsruhe, Germany

    Stefan A. Oelmeier & Jürgen Hubbuch

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  1. Andrea Schaller
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  2. Natalie K. Connors
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  3. Mirjana Dimitrijev Dwyer
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  4. Stefan A. Oelmeier
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  6. Anton P. J. Middelberg
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Correspondence to Natalie K. Connors.

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Schaller, A., Connors, N.K., Dwyer, M.D. et al. Computational study of elements of stability of a four-helix bundle protein biosurfactant. J Comput Aided Mol Des 29, 47–58 (2015). https://doi.org/10.1007/s10822-014-9803-6

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