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Synthesis of an antiviral drug precursor from chitin using a saprophyte as a whole-cell catalyst

Matthias G Steiger14, Astrid R Mach-Aigner1, Rita Gorsche1, Erwin E Rosenberg2, Marko D Mihovilovic3 and Robert L Mach1*

Author Affiliations

1 Gene Technology and Applied Biochemistry, Institute of Chemical Engineering, Vienna University of Technology, Gumpendorfer Str. 1a, A-1060 Wien, Austria

2 Instrumental Analytical Chemistry, Institute of Chemical Technologies and Analytics, Vienna University of Technology, Getreidemarkt 9/164, A-1060 Wien, Austria

3 Institute of Applied Synthetic Chemistry, Vienna University of Technology, Getreidemarkt 9/163, A-1060 Wien, Austria

4 Austrian Centre of Industrial Biotechnology, Muthgasse 107, 1190 Vienna, Austria

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Microbial Cell Factories 2011, 10:102  doi:10.1186/1475-2859-10-102

Published: 5 December 2011

Abstract

Background

Recent incidents, such as the SARS and influenza epidemics, have highlighted the need for readily available antiviral drugs. One important precursor currently used for the production of Relenza, an antiviral product from GlaxoSmithKline, is N-acetylneuraminic acid (NeuNAc). This substance has a considerably high market price despite efforts to develop cost-reducing (biotechnological) production processes. Hypocrea jecorina (Trichoderma reesei) is a saprophyte noted for its abundant secretion of hydrolytic enzymes and its potential to degrade chitin to its monomer N-acetylglucosamine (GlcNAc). Chitin is considered the second most abundant biomass available on earth and therefore an attractive raw material.

Results

In this study, we introduced two enzymes from bacterial origin into Hypocrea, which convert GlcNAc into NeuNAc via N-acetylmannosamine. This enabled the fungus to produce NeuNAc from the cheap starting material chitin in liquid culture. Furthermore, we expressed the two recombinant enzymes as GST-fusion proteins and developed an enzyme assay for monitoring their enzymatic functionality. Finally, we demonstrated that Hypocrea does not metabolize NeuNAc and that no NeuNAc-uptake by the fungus occurs, which are important prerequisites for a potential production strategy.

Conclusions

This study is a proof of concept for the possibility to engineer in a filamentous fungus a bacterial enzyme cascade, which is fully functional. Furthermore, it provides the basis for the development of a process for NeuNAc production as well as a general prospective design for production processes that use saprophytes as whole-cell catalysts.