Journal of Bacteriology, November 1998, p. 5547-5558, Vol. 180, No. 21
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Department of Microbiology, University of Illinois, Urbana, Illinois 61801
Received 15 May 1998/Accepted 17 August 1998
The first molecular and genetic characterization of a biochemical
pathway for oxidation of the reduced phosphorus (P) compounds phosphite
and hypophosphite is reported. The pathway was identified in
Pseudomonas stutzeri WM88, which was chosen for detailed
studies from a group of organisms isolated based on their ability to
oxidize hypophosphite (+1 valence) and phosphite (+3 valence) to
phosphate (+5 valence). The genes required for oxidation of both
compounds by P. stutzeri WM88 were cloned on a single ca.
30-kbp DNA fragment by screening for expression in Escherichia
coli and Pseudomonas aeruginosa. Two lines of
evidence suggest that hypophosphite is oxidized to phosphate via a
phosphite intermediate. First, plasmid subclones that conferred
oxidation of phosphite, but not hypophosphite, upon heterologous hosts
were readily obtained. All plasmid subclones that failed to confer
phosphite oxidation also failed to confer hypophosphite oxidation. No
subclones that conferred only hypophosphite expression were obtained.
Second, various deletion derivatives of the cloned genes were made in
vitro and recombined onto the chromosome of P. stutzeri
WM88. Two phenotypes were displayed by individual mutants. Mutants with
the region encoding phosphite oxidation deleted (based upon the
subcloning results) lost the ability to oxidize either phosphite or
hypophosphite. Mutants with the region encoding hypophosphite oxidation
deleted lost only the ability to oxidize hypophosphite. The phenotypes
displayed by these mutants also demonstrate that the cloned genes are
responsible for the P oxidation phenotypes displayed by the original
P. stutzeri WM88 isolate. The DNA sequences of the minimal
regions implicated in oxidation of each compound were determined. The
region required for oxidation of phosphite to phosphate putatively
encodes a binding-protein-dependent phosphite transporter, an
NAD+-dependent phosphite dehydrogenase, and a
transcriptional activator of the lysR family. The region
required for oxidation of hypophosphite to phosphite putatively encodes
a binding-protein-dependent hypophosphite transporter and an
-ketoglutarate-dependent hypophosphite dioxygenase. The finding of
genes dedicated to oxidation of reduced P compounds provides further
evidence that a redox cycle for P may be important in the metabolism of
this essential, and often growth-limiting, nutrient.
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