Abstract
BackgroundPachysolen tannophilus is a non-conventional yeast, which can metabolize many of the carbon sources found in low cost feedstocks including glycerol and xylose. The xylose utilisation pathways have been extensively studied in this organism. However, the mechanism behind glycerol metabolism is poorly understood. Using the recently published genome sequence of P. tannophilus CBS4044, we searched for genes with functions in glycerol transport and metabolism by performing a BLAST search using the sequences of the relevant genes from Saccharomyces cerevisiae as queries.ResultsQuantitative real-time PCR was performed to unveil the expression patterns of these genes during growth of P. tannophilus on glycerol and glucose as sole carbon sources. The genes predicted to be involved in glycerol transport in P. tannophilus were expressed in S. cerevisiae to validate their function. The S. cerevisiae strains transformed with heterologous genes showed improved growth and glycerol consumption rates with glycerol as the sole carbon source.ConclusionsP. tannophilus has characteristics relevant for a microbial cell factory to be applied in a biorefinery setting, i.e. its ability to utilise the carbon sources such as xylose and glycerol. However, the strain is not currently amenable to genetic modification and transformation. Heterologous expression of the glycerol transporters from P. tannophilus, which has a relatively high growth rate on glycerol, could be used as an approach for improving the efficiency of glycerol assimilation in other well characterized and applied cell factories such as S. cerevisiae.
Highlights
Pachysolen tannophilus is a non-conventional yeast, which can metabolize many of the carbon sources found in low cost feedstocks including glycerol and xylose
Identification of genes in glycerol metabolism and analysis of glycerol transporters in P. tannophilus In order to find homologous genes related to glycerol transport and metabolism in P. tannophilus, the genome sequence was searched by BLAST with the sequences of genes known to have these functions in S. cerevisiae as queries
In D. hansenii, it was shown that the active glycerol transport system was constitutively expressed and not subject to glucose repression [27]. In agreement with these experiments, we showed that the glycerol symporter genes PtSTL1 and PtSTL2 in P. tannophilus are constitutively expressed on glycerol and glucose based on quantitative real-time PCR (qPCR) expression analysis
Summary
Pachysolen tannophilus is a non-conventional yeast, which can metabolize many of the carbon sources found in low cost feedstocks including glycerol and xylose. P. tannophilus has been shown to be capable of converting crude glycerol to ethanol under microaerobic conditions [2] This ability is interesting since glycerol, a by-product of biodiesel production, has been considered as a potential alternative carbon source for industrial bioprocesses due to the recent dramatic increase in production of biodiesel. In S. cerevisiae, glycerol enters the cell by two different mechanisms: a low affinity transport system (facilitated diffusion) and a high affinity proton symport system (active transport) [7]. A screen for genes encoding membrane proteins involved in glycerol assimilation in S. cerevisiae identified a gene, STL1, involved in active glycerol uptake. It has been shown that in Candida albicans glycerol was actively transported into the cells by a proton symporter encoded by the C. albicans STL1 [13]. It has been reported that S. cerevisiae strains harboring the STL1 gene from D. hansenii slightly improved their growth and doubling times on glycerol [14]
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