Schempp Florence Miramella, Hofmann Katharina Elisabeth, Mi Jia, Kirchner Ferdinand, Meffert Annika, Schewe Hendrik, Schrader Jens, Buchhaupt Markus
DECHEMA-Forschungsinstitut, Industrial Biotechnology, Theodor-Heuss-Allee 25, 60486, Frankfurt am Main, Germany.
Faculty Biological Sciences, Goethe University Frankfurt, Max-von-Laue-Str. 9, 60438, Frankfurt am Main, Germany.
Appl Microbiol Biotechnol. 2020 Jun;104(12):5519-5533. doi: 10.1007/s00253-020-10566-3. Epub 2020 Apr 16.
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.
单萜类化合物广泛应用于工业领域，例如作为药物、香料和香精组合物以及农业中的活性成分。一些单萜类化合物具有严重的毒性作用，这使得它们成为生物技术生产过程中具有挑战性的化合物。恶臭假单胞菌的一些菌株对包括单萜类化合物在内的溶剂表现出固有的极高耐受性。了解其潜在因素有助于创建适合从头生产或转化单萜类化合物的菌株。此外，关于耐受机制的知识可以更深入地了解细菌如何对抗含单萜类化合物的药物，如茶树油。在这项工作中，使用选定的单萜类化合物超耐受突变体研究了恶臭假单胞菌GS1的抗性机制。大多数突变发生在流出泵启动子区域或相关转录因子中。令人惊讶的是，对于测试的单萜类醇、酮和醚，高流出泵表达增加了对单萜类化合物的耐受性，但降低了对香叶酸的耐受性。然而，通过流出泵组件中的突变可以提高对香叶酸的耐受性。还发现，单萜类化合物耐受性的增加会抵消有效的生物转化能力，这表明在生产菌株开发中需要进行微调且基于知识的耐受性改进。要点：• 单萜类化合物耐受性的改变主要与流出泵活性的改变有关。• 对香叶酸耐受性的增加令人惊讶地是由输出活性降低引起的。• 输出活性的降低对生物技术转化可能有益。
