Structural and Functional Analysis of the Phosphonoacetate Hydrolase ( phnA ) Gene Region in Pseudomonas fluorescens 23F

Abstract

The Pseudomonas fluorescens 23F phosphonoacetate hydrolase gene (phnA) encodes a novel carbon-phosphorus bond cleavage enzyme whose expression is independent of the phosphate status of the cell. Analysis of the regions adjacent to the phosphonoacetate hydrolase structural gene (phnA) indicated the presence of five open reading frames (ORFs). These include one (phnR) whose putative product shows high levels of homology to the LysR family of positive transcriptional regulators. Its presence was shown to be necessary for induction of the hydrolase activity. 2-Phosphonopropionate was found to be an inducer (and poor substrate) for phosphonoacetate hydrolase. Unlike phosphonoacetate, which is also an inducer of phosphonoacetate hydrolase, entry of 2-phosphonopropionate into cells appeared to be dependent on the presence of a gene (phnB) that lies immediately downstream of phnA and whose putative product shows homology to the glycerol-3-phosphate transporter. RNA analysis revealed transcripts for the phnAB and phnR operons, which are transcribed divergently; the resulting mRNAs overlapped by 29 nucleotide bases at their 5 ends. Transcripts of phnAB were detected only in cells grown in the presence of phosphonoacetate, whereas transcripts of phnR were observed in cells grown under both induced and uninduced conditions. The expression of three additional genes found in the phnA region did not appear necessary for the degradation of phosphonoacetate and 2-phosphonopropionate by either Pseudomonas putida or Escherichia coli cells.Organophosphonates are a group of compounds of biogenic and xenobiotic origins that are characterized by possession of a direct carbon-phosphorus bond. Two different routes of C-P bond cleavage, each inducible only under conditions of phosphorus limitation, have been demonstrated when such molecules serve as the sole phosphorus source for microbial growth. "C-P lyase" is the trivial name given to an enzyme complex which catalyzes the cleavage of the C-P bond of both substituted and unsubstituted phosphonates by a mechanism which may involve redox or radical chemistry (27). By contrast, phosphonoacetaldehyde hydrolase ("phosphonatase" [EC 3.11.1.1]) is active in vitro and hydrolytically cleaves only the substituted C-P bond of phosphonoacetaldehyde to yield acetaldehyde and P i (10). Since bacterial cleavage of the C-P bond by both C-P lyase(s) and phosphonatase is under control of the pho regulon (10) and hence occurs only under conditions of P i limitation, organophosphonates generally fail to serve as sources of carbon for microbial growth because the excess of P i released during the catabolism of their carbon skeletons serves to repress and/or inhibit their further mineralization. Genetic analysis of C-P bond cleavage by the C-P lyase and phosphonatase pathways has been reported (10,18,26), and the phosphonatase gene of Pseudomonas aeruginosa has been characterized (5).We have recently reported the purification (15) and molecular cloning (14) of phosphonoacetate (PA) hydrolase (EC 3.11.1.2), ...

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“…The majority of LTTR-encoding genes are monocistronic and are transcribed divergently from the genes under their control (18,25,30). In contrast, the virR gene is cotranscribed with four other genes located downstream from virR.…”

Utilization of Glyphosate as Phosphate Source: Biochemistry and Genetics of Bacterial Carbon-Phosphorus Lyase

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Comparative genomics analyses on EPS biosynthesis genes required for floc formation of Zoogloea resiniphila and other activated sludge bacteria

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