Comparative transcriptomics and proteomics of p-hydroxybenzoate producing Pseudomonas putida S12: novel responses and implications for strain improvement

Abstract

A transcriptomics and proteomics approach was employed to study the expression changes associated with p-hydroxybenzoate production by the engineered Pseudomonas putida strain S12palB1. To establish p-hydroxybenzoate production, phenylalanine-tyrosine ammonia lyase (pal/tal) was introduced to connect the tyrosine biosynthetic and p-coumarate degradation pathways. In agreement with the efficient p-hydroxybenzoate production, the tyrosine biosynthetic and p-coumarate catabolic pathways were upregulated. Also many transporters were differentially expressed, one of which—a previously uncharacterized multidrug efflux transporter with locus tags PP1271-PP1273—was found to be associated with p-hydroxybenzoate export. In addition to tyrosine biosynthesis, also tyrosine degradative pathways were upregulated. Eliminating the most prominent of these resulted in a 22% p-hydroxybenzoate yield improvement. Remarkably, the upregulation of genes contributing to p-hydroxybenzoate formation was much higher in glucose than in glycerol-cultured cells.Electronic supplementary materialThe online version of this article (doi:10.1007/s00253-010-2626-z) contains supplementary material, which is available to authorized users.

Efflux systems in bacteria and their metabolic engineering applications

“…It is assumed that efflux systems are facilitating the export of the aliphatic products in these situations, yet few reports have directly examined this hypothesis. In one example, production of p-hydroxybenzoic acid (PHB) from glucose in P. putida S12 results in the up-regulation of the tripartite efflux system PP_1271-73, which is homologous to emrAB - tolC in E. coli (Verhoef et al, 2010). Disrupting PP_1271-73 in a PHB production strain decreases the final titer of PHB almost 2-fold, along with a concomitant decrease in cell growth, however, overexpressing the tripartite system did not increase titers (Verhoef et al, 2010).…”

Anodic respiration of Pseudomonas putida KT2440 in a stirred-tank bioreactor

“…So far pHBA has been produced under aerobic conditions from glucose in recombinant strains of Escherichia coli [15], Saccharomyces cerevisiae [16,17] and Pseudomonas putida [18,19]. Maximal pHBA yields of 49 mmolC pHBA molC -1 glucose were achieved with P. putida in a chemostat cultivation [19].…”

Redundancy in putrescine catabolism in solvent tolerant Pseudomonas putida S12

“…Recently, novel indications for such a role of polyamines, particularly putrescine, were found in a transcriptome analysis of a p-hydroxybenzoate producing P. putida S12 strain. This study revealed a number of genes presumably involved in putrescine transport and metabolism that were differentially expressed in this strain compared to a non-producing control strain (Verhoef et al, 2010). Also in other studies, polyamine-associated genes were found to be differentially expressed in solvent-exposed P. putida strains (Duque et al, 2007;Volkers et al, 2009).…”