Enhancement of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) accumulation in Arxula adeninivorans by stabilization of production

Mateusz Biernacki,Thomas Roick,Marek Marzec,Kim Baronian,Gotthard Kunze,Rüdiger Bode,Reinhard Pätz

doi:10.1186/s12934-017-0751-4

Abstract

BackgroundIn recent years the production of biobased biodegradable plastics has been of interest of researchers partly due to the accumulation of non-biodegradable plastics in the environment and to the opportunity for new applications. Commonly investigated are the polyhydroxyalkanoates (PHAs) poly(hydroxybutyrate) and poly(hydroxybutyrate-co-hydroxyvalerate) (PHB-V). The latter has the advantage of being tougher and less brittle. The production of these polymers in bacteria is well established but production in yeast may have some advantages, e.g. the ability to use a broad spectrum of industrial by-products as a carbon sources.ResultsIn this study we increased the synthesis of PHB-V in the non-conventional yeast Arxula adeninivorans by stabilization of polymer accumulation via genetic modification and optimization of culture conditions. An A. adeninivorans strain with overexpressed PHA pathway genes for β-ketothiolase, acetoacetyl-CoA reductase, PHAs synthase and the phasin gene was able to accumulate an unexpectedly high level of polymer. It was found that an optimized strain cultivated in a shaking incubator is able to produce up to 52.1% of the DCW of PHB-V (10.8 g L−1) with 12.3%mol of PHV fraction. Although further optimization of cultivation conditions in a fed-batch bioreactor led to lower polymer content (15.3% of the DCW of PHB-V), the PHV fraction and total polymer level increased to 23.1%mol and 11.6 g L−1 respectively. Additionally, analysis of the product revealed that the polymer has a very low average molecular mass and unexpected melting and glass transition temperatures.ConclusionsThis study indicates a potential of use for the non-conventional yeast, A. adeninivorans, as an efficient producer of polyhydroxyalkanoates.

Highlights

In recent years the production of biobased biodegradable plastics has been of interest of researchers partly due to the accumulation of non-biodegradable plastics in the environment and to the opportunity for new applications
The entire industrial production of PHB-V is currently based on bacteria, which may be associated with certain problems such as phage contamination of Escherichia coli process [10] or possible presence of lipopolysaccharides in the product, which excludes its use in medical applications [11]
PHA shaking flask screening Overexpression of ß-ketothiolase gene and acetoacetyl-CoA reductase and PHA synthase genes from C. necator H16 in A. adeninivorans led to synthesis of PHA by this yeast

Summary

Introduction

In recent years the production of biobased biodegradable plastics has been of interest of researchers partly due to the accumulation of non-biodegradable plastics in the environment and to the opportunity for new applications. Synthesis of PHB-V is carried out by at least three enzymes: β-ketothiolase, (R)-specific NADPH-dependent acetoacetyl-CoA reductase and PHA synthase [6]. This pathway has been primarily investigated in bacteria, e.g. Cupriavidus necator for PHB-V production with range of accumulation level between 50 and 90% of DCW containing up to 24% PHV [1, 7]. Phasins are a group of low-molecular-weight proteins with amphiphilic properties In bacteria, where these proteins were originally found, they play a regulatory and stability role during PHA synthesis and cell division [17]. C. necator phasin PhaP1 gene, unexpectedly decreased cells stress when overexpressed in a non-phasin strain of E. coli [18]

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