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A Pseudomonas putida bioreporter for the detection of enzymes active on 2-alkyl-4(1H)-quinolone signalling molecules.

Abstract：

:The quorum sensing signalling molecules 2-heptyl-3-hydroxy-4(1H)-quinolone, termed the "Pseudomonas quinolone signal" (PQS), and 2-heptyl-4(1H)-quinolone (HHQ) play an important role in the control of virulence gene expression in Pseudomonas aeruginosa. To construct a bioreporter for the specific and sensitive detection of these compounds, a plasmid with the pqsR gene encoding the PQS- and HHQ-responsive transcriptional regulator PqsR, and with the PqsR-controlled pqsA promoter fused to the lacZ gene, was established in Pseudomonas putida KT2440. The bioreporter responds to HHQ and PQS at concentrations in the range of 0.1-10 and 0.01-5 μM, respectively, with EC(50) values of 1.50 ± 0.25 μM for HHQ and 0.15 ± 0.02 μM for PQS. 2,4-Dihydroxyquinoline, a metabolite produced abundantly by P. aeruginosa, did not elicit an increase in reporter enzyme activity. To test whether the bioreporter can be used for the detection of enzymes active on AQ signalling molecules, the hodC gene coding for 2-methyl-3-hydroxy-4(1H)-quinolone 2,4-dioxygenase was expressed in the reporter strain. This dioxygenase catalyses the cleavage of PQS, albeit with very low activity. The response of the bioreporter to PQS was significantly quenched by co-expression of the hodC gene, and HPLC analysis of culture extracts verified that the PQS levels decreased during cultivation. The bioreporter can be applied to screen for AQ-converting enzymes, which will be useful tools to interfere with quorum sensing and thus virulence in P. aeruginosa.

journal_name

Appl Microbiol Biotechnol

journal_title

Applied microbiology and biotechnology

authors

Müller C,Fetzner S

doi

10.1007/s00253-012-4236-4

subject

Has Abstract

pub_date

2013-01-01 00:00:00

pages

751-60

issue

2

eissn

0175-7598

issn

1432-0614

journal_volume

97

pub_type

杂志文章

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