Abstract
Recombinant strains of Wautersia eutropha expressing an artificial polyhydroxyalkanoate (PHA) biosynthesis operon under the control of different native promoters linked to polyhydroxybutyrate (PHB) (Pphb), acetoin (PacoE, PacoD, and PacoX) or pyruvate (PpdhE) metabolism were constructed and tested. The promoters were representative either of the enterobacterial σ70 (Pphb, PacoE, and PpdhE)- or σ54 (PacoD and PacoX)-dependent promoters. To obtain polymers consisting of C4–C12 monomer units, an artificial operon consisting of the PHA synthase gene from Pseudomonas sp. 61-3 (phaC1 Ps) tandemly linked to the W. eutropha genes encoding β-ketothiolase (phbA We) and nicotinamide adenine dinucleotide phosphate dependent acetoacetyl-coenzyme A (CoA) reductase (phbB We) was constructed. All recombinant strains produced PHA, indicating that the PHA biosynthesis genes were expressed under the control of the different promoters. Cell growth and PHA synthesis on MS medium complemented with gluconate or octanoate, and different concentrations of acetoin (0, 0.15, and 0.3%) clearly differed among the recombinant strains. While the PacoD and PacoX promoters mediated only low PHA yields (<1%) in the presence of the inducer acetoin, the remaining promoters—independent of the addition of acetoin—resulted in the production of PHA polymers with high 3HB fractions (90–100 mol%) and with high 3HO contents (70–86 mol%) from gluconate and octanoate, respectively. Interestingly, on octanoate-MS medium with 0.15% acetoin, the PacoE promoter mediated the synthesis of PHA with a relatively high 3HB fraction (48 mol%). While PHAs with high 3HB contents were obtained, the overall PHA product yields were low (<10%); thus, their potential application for further commercial exploitation appears limited.
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Acknowledgement
We thank Procter & Gamble (Cincinnati, USA) for supporting this work and providing us with the pBSEB50 and pBBR1MCS-2/Pphb constructs.
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Delamarre, S.C., Batt, C.A. Comparative study of promoters for the production of polyhydroxyalkanoates in recombinant strains of Wautersia eutropha . Appl Microbiol Biotechnol 71, 668–679 (2006). https://doi.org/10.1007/s00253-005-0217-1
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DOI: https://doi.org/10.1007/s00253-005-0217-1