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Article

Key Factors for a One-Pot Enzyme Cascade Synthesis of High Molecular Weight Hyaluronic Acid

1
Laboratory for Biomaterials, Institute of Biotechnology and Helmholtz-Institute for Biomedical Engineering, RWTH Aachen University, Pauwelsstraße 20, 52074 Aachen, Germany
2
Research and Development Department, GALAB Laboratories GmbH, Am Schleusengraben 7, 21029 Hamburg, Germany
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2019, 20(22), 5664; https://doi.org/10.3390/ijms20225664
Received: 24 October 2019 / Revised: 11 November 2019 / Accepted: 11 November 2019 / Published: 12 November 2019
(This article belongs to the Special Issue Molecular Biocatalysis 2.0)
In the last decades, interest in medical or cosmetic applications of hyaluronic acid (HA) has increased. Size and dispersity are key characteristics of biological function. In contrast to extraction from animal tissue or bacterial fermentation, enzymatic in vitro synthesis is the choice to produce defined HA. Here we present a one-pot enzyme cascade with six enzymes for the synthesis of HA from the cheap monosaccharides glucuronic acid (GlcA) and N-acetylglucosamine (GlcNAc). The combination of two enzyme modules, providing the precursors UDP–GlcA and UDP–GlcNAc, respectively, with hyaluronan synthase from Pasteurella multocida (PmHAS), was optimized to meet the kinetic requirements of PmHAS for high HA productivity and molecular weight. The Mg2+ concentration and the pH value were found as key factors. The HA product can be tailored by different conditions: 25 mM Mg2+ and 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid (HEPES)-NaOH pH 8 result into an HA product with high Mw HA (1.55 MDa) and low dispersity (1.05). Whereas with 15 mM Mg2+ and HEPES–NaOH pH 8.5, we reached the highest HA concentration (2.7 g/L) with a yield of 86.3%. Our comprehensive data set lays the basis for larger scale enzymatic HA synthesis. View Full-Text
Keywords: hyaluronic acid; in vitro synthesis; one-pot multi-enzyme; optimization; enzyme cascade hyaluronic acid; in vitro synthesis; one-pot multi-enzyme; optimization; enzyme cascade
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MDPI and ACS Style

Gottschalk, J.; Zaun, H.; Eisele, A.; Kuballa, J.; Elling, L. Key Factors for a One-Pot Enzyme Cascade Synthesis of High Molecular Weight Hyaluronic Acid. Int. J. Mol. Sci. 2019, 20, 5664. https://doi.org/10.3390/ijms20225664

AMA Style

Gottschalk J, Zaun H, Eisele A, Kuballa J, Elling L. Key Factors for a One-Pot Enzyme Cascade Synthesis of High Molecular Weight Hyaluronic Acid. International Journal of Molecular Sciences. 2019; 20(22):5664. https://doi.org/10.3390/ijms20225664

Chicago/Turabian Style

Gottschalk, Johannes, Henning Zaun, Anna Eisele, Jürgen Kuballa, and Lothar Elling. 2019. "Key Factors for a One-Pot Enzyme Cascade Synthesis of High Molecular Weight Hyaluronic Acid" International Journal of Molecular Sciences 20, no. 22: 5664. https://doi.org/10.3390/ijms20225664

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