Enhancement of acetoin production in Candida glabrata by in silico-aided metabolic engineering.

Shubo Li,Xiang Gao,Liming Liu,Jian Chen,Nan Xu

doi:10.1186/1475-2859-13-55

Shubo Li, Xiang Gao + Show 3 more

Open Access

https://doi.org/10.1186/1475-2859-13-55

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Abstract

BackgroundAcetoin is a promising chemical compound that can potentially serve as a high value-added platform for a broad range of applications. Many industrial biotechnological processes are moving towards the use of yeast as a platform. The multi-auxotrophic yeast, Candida glabrata, can accumulate a large amount of pyruvate, but produces only trace amounts of acetoin. Here, we attempted to engineer C. glabrata to redirect the carbon flux of pyruvate to increase acetoin production.ResultsBased on an in silico strategy, a synthetic, composite metabolic pathway involving two distinct enzymes, acetolactate synthase (ALS) and acetolactate decarboxylase (ALDC), was constructed, leading to the accumulation of acetoin in C. glabrata. Further genetic modifications were introduced to increase the carbon flux of the heterologous pathway, increasing the production of acetoin to 2.08 g/L. Additionally, nicotinic acid was employed to regulate the intracellular NADH level, and a higher production of acetoin (3.67 g/L) was obtained at the expense of 2,3-butanediol production under conditions of a lower NADH/NAD+ ratio.ConclusionWith the aid of in silico metabolic engineering and cofactor engineering, C. glabrata was designed and constructed to improve acetoin production.

Highlights

Acetoin is a promising chemical compound that can potentially serve as a high value-added platform for a broad range of applications
To realize acetoin accumulation in C. glabrata, its metabolic capacity was engineered by introducing a heterologous pathway in the cytosol, and evaluated by in silico simulation
According to the approach previously described [19,20,21], the reactions representing the enzymatic activities of αacetolactate synthetase (ALS) and α-acetolactate decarboxylase (ALDC) were added into the model i NX804 prior to flux balance analysis, using acetoin as the objective equation by the FBA algorithm (Additional file 1a)

Summary

Introduction

Acetoin is a promising chemical compound that can potentially serve as a high value-added platform for a broad range of applications. The multi-auxotrophic yeast, Candida glabrata, can accumulate a large amount of pyruvate, but produces only trace amounts of acetoin. As a member of the C4-dicarboxylic acid family, acetoin was defined as one of the potential top 30 sugar-derived chemical building blocks by the U.S Department of Energy [1], and has drawn much interest because it could serve as a high value-added platform for the food, flavor, cosmetics, pharmaceutical, and chemical industries [2,3]. Three methods are used to produce acetoin: microbial fermentation, enzymatic conversion, and chemical synthesis [4]. The biological platform for microbial acetoin production needs to be further refined and improved

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