Inducible expression of choline sulfatase and its regulator BetR in Pseudomonas sp. ATCC19151
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ORIGINAL PAPER
Inducible expression of choline sulfatase and its regulator BetR in Pseudomonas sp. ATCC19151 Branko Jovcic · Vittorio Venturi · Ljubisa Topisirovic · Milan Kojic
Received: 31 August 2010 / Revised: 11 February 2011 / Accepted: 17 February 2011 / Published online: 3 March 2011 © Springer-Verlag 2011
Abstract Pseudomonas sp. strain ATCC19151 is a natural isolate from sewage with the ability to degrade detergents. Genes encoding potential choline sulfatase (betC), substrate-binding ABC transporter protein (betD), sulfate transporter (betE), and divergent putative transcriptional regulator (betR) were cloned and characterized from strain ATCC19151. In silico analysis revealed that (1) the BetC protein belongs to alkPPc superfamily and shares CXPXR sequence with the cysteine sulfatases of group I, (2) BetR belongs to the LysR family of transcriptional regulators, (3) BetD is part of the PBPb superfamily of periplasmic and membrane-associated proteins, and (4) BetE is a permease and contains STAS domain. Insertional mutagenesis and genetic complementation show that betC gene encodes a functional choline sulfatase. Analysis of the betC (PbetC) and betR (PbetR) promoters revealed that they are inducible. BetR activates betC and betR transcription in the presence of choline sulfate, whilst in the absence of choline sulfate, BetR represses its own transcription. It was further established that BetR directly binds to betC–betR intergenic region in vitro, with higher aYnity in the presence of choline sulfate as cofactor. Transcription of betC and betR was not induced in the presence of high concentration of NaCl.
Communicated by Jorge Membrillo-Hernandez. B. Jovcic · L. Topisirovic · M. Kojic (&) Institute of Molecular Genetics and Genetic Engineering, Vojvode Stepe 444a, P.O. Box 23, 11010 Belgrade, Serbia e-mail: [email protected] V. Venturi International Centre for Genetic Engineering and Biotechnology, AREA Science Park, Padriciano 99, 34149 Trieste, Italy
Keywords Pseudomonas · Choline sulfatase · Transcriptional regulation of gene expression · Osmoprotection
Introduction Bacteria cope with the osmotic stress by accumulation of compatible solutes, compounds that balance turgor pressure. Compatible solutes can be accumulated to high intracellular concentrations without interfering with physiological processes. Glycine betaine for example is a widely used compatible solute, which can be synthesized de novo or accumulated from external sources (Miller and Wood 1996). Glycine betaine is commonly synthesized from choline, and several enzymatic systems have been reported to catalyze such a conversion. Two-step transformation of choline to betaine is mediated by means of subsequent betaine aldehyde dehydrogenase and choline dehydrogenase activities in Escherichia coli (Lamark et al. 1991), Halomonas elongata (Canovas et al. 2000) and Sinorhizobium meliloti (Pocard et al. 1997). Some bacteria and fungi can produce choline from choline-O-sulfate (COS) by means of a choline sulfatase activity (Lucas et al. 1972;
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