Inorganic Polyphosphate in Vibrio cholerae : Genetic, Biochemical, and Physiologic Features

Abstract

Vibrio cholerae O1, biotype El Tor, accumulates inorganic polyphosphate (poly P) principally as large clusters of granules. Poly P kinase (PPK), the enzyme that synthesizes poly P from ATP, is encoded by the ppk gene, which has been cloned from V. cholerae, overexpressed, and knocked out by insertion-deletion mutagenesis. The predicted amino acid sequence of PPK is 701 residues (81.6 kDa), with 64% identity to that of Escherichia coli, which it resembles biochemically. As in E. coli, ppk is part of an operon with ppx, the gene that encodes exopolyphosphatase (PPX). However, unlike in E. coli, PPX activity was not detected in cell extracts of wild-type V. cholerae. The ppk null mutant of V. cholerae has diminished adaptation to high concentrations of calcium in the medium as well as motility and abiotic surface attachment.Inorganic polyphosphate poly P is a linear polymer of up to hundreds of orthophosphate (P i ) residues linked by high-energy phosphoanhydride bonds. Among known functions, poly P can serve as a substitute for ATP in kinase reactions, a P i reservoir, and a chelator of divalent metals (9). Poly P is ubiquitous in nature, having been found in all organisms examined (15), yet little is known about its physiological roles (14).Several poly P-metabolizing enzymes have been purified, and the genes encoding them have been cloned (14, 28). The enzyme primarily responsible for poly P synthesis in Escherichia coli is poly P kinase (PPK), which catalyzes the polymerization of the ␥ phosphate of ATP into a poly P chain (1). Poly P can be hydrolyzed to P i by an exopolyphosphatase (PPX) (3). In E. coli, the encoding genes, ppk and ppx, respectively, form an operon. The inability to accumulate poly P upon deletion of this operon or upon the overproduction of PPX has produced several striking phenotypes in E. coli (6,20,25): decreased long-term survival in stationary phase; increased sensitivity to oxidative, osmotic, and thermal stresses; and defects in adaptive growth in minimal medium after a shift from rich medium. These phenotypes are likely due to the decreased expression of the rpoS gene, which encodes the principal stationary-phase sigma factor, S , or RpoS (25). These and related results (4) suggest that poly P is an effector signal for responses to acute stringencies and adaptations in the stationary phase.Recently available genome sequences have revealed that PPK is highly conserved in many bacterial species, including some important pathogens (26). This also implies that PPK and/or poly P has fundamental physiological roles in bacteria. The ppk knockout mutant of Pseudomonas aeruginosa PAO1 shows a dramatic deficiency in motility, both flagellar and pilus mediated, an inability to form biofilms, and a loss of virulence (22,23,24). ppk null mutants of several other pathogens and of E. coli also exhibit reduced motility and reduced abiotic surface attachment (22,24).Vibrio spp. are among the most common microorganisms in environmental surface waters, such as lakes and rivers. Vibrio cholerae O1 is an...

Inorganic polyphosphate in Bacillus cereus : Motility, biofilm formation, and sporulation

“…Impaired Motility in the Mutants. Deletion of ppk1 in many Gramnegative strains, such as E. coli, P. aeruginosa, and Vibrio cholerae, impairs their motility (14,22). In B. cereus, flagellar motility observed as swimming and swarming was impaired not only in the ⌬ppk mutant but even more so in the ⌬ppx mutant, despite a massive accumulation of poly-P in this mutant (Fig.…”

“…(29), Shigella flexneri (29), V. cholerae (22), and Helicobacter pylori (3), that of B. cereus is the most distinctive. The PPK of B. cereus has a low pI, fails to adsorb to anion exchangers, and is stimulated 10-fold in activity in lysates by the presence of poly-P.…”

Copper tolerance of the thermoacidophilic archaeon Sulfolobus metallicus: possible role of polyphosphate metabolism

“…) for another micro-organism accumulating polyP granules -Vibrio cholerae during the exponential phase of growth with a rich medium (Ogawa et al, 2000) -and are closer to the 400 nmol (mg protein) 21 accumulated by A. ferrooxidans (Alvarez & Jerez, 2004). It is possible that the polyP accumulations observed in these crenarchaea contribute not only to their energy supply but also to their enhanced survival and stress resistance in the presence of metals, as suggested for bacteria (Keasling, 1997;Alvarez & Jerez, 2004).…”

Environmental phosphate differentially affects virulence phenotypes of uropathogenicEscherichia coliisolates causative of prostatitis

“…12,13 Inorganic polyphosphate (polyP) was shown to be critical for attributes such as motility, quorum sensing, biofilm formation, resistance to oxidative, osmotic, heat, nutritional and alkaline stress, and stationary-phase survival in several microorganisms. [14][15][16][17][18][19][20][21][22][23] PolyP is a linear chain of tens to many hundreds of phosphate residues linked by high-energy phosphoanhydride bonds. 14,24 PolyP is usually accumulated during exponential phase of growth, and degraded at the beginning of stationary phase.…”