Investigation of monoterpenoid resistance mechanisms in Pseudomonas putida and their consequences for biotransformations

F.M. Schempp, K.E. Hofmann, J.Mi, F. Kirchner, A. Meffert, H. Schewe, J. Schrader, M. Buchhaupt

Monoterpenoids are widely used in industrial applications, e.g. as active ingredients in pharmaceuticals, in flavor and fragrance compositions, and in agriculture. Severe toxic effects are known for some monoterpenoids making them challenging compounds for biotechnological production processes. Some strains of the bacterium Pseudomonas putida show an inherent extraordinarily high tolerance towards solvents including monoterpenoids. An understanding of the underlying factors can help to create suitable strains for monoterpenoids de novo production or conversion. In addition, knowledge about tolerance mechanisms could allow a deeper insight into how bacteria can oppose monoterpenoid containing drugs, like tea tree oil. Within this work, the resistance mechanisms of P. putida GS1 were investigated using selected monoterpenoid-hypertolerant mutants. Most of the mutations were found in efflux pump promoter regions or associated transcription factors. Surprisingly, while for the tested monoterpenoid alcohols, ketone, and ether high efflux pump expression increased monoterpenoid tolerance, it reduced the tolerance against geranic acid. However, an increase of geranic acid tolerance could be gained by a mutation in an efflux pump component. It was also found that increased monoterpenoid tolerance can counteract efficient biotransformation ability, indicating the need for a fine-tuned and knowledge-based tolerance improvement for production strain development.Key points• Altered monoterpenoid tolerance mainly related to altered activity of efflux pumps.• Increased tolerance to geranic acid surprisingly caused by decreased export activity. • Reduction of export activity can be beneficial for biotechnological conversions.

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