Antibiotic and biosurfactant properties of cyclic lipopeptides produced by fluorescent Pseudomonas spp. from the sugar beet rhizosphere.

Cyclic lipopeptides (CLPs) with antibiotic and biosurfactant properties are produced by a number of soil bacteria, including fluorescent Pseudomonas spp. To provide new and efficient strains for the biological control of root-pathogenic fungi in agricultural crops, we isolated approximately 600 fluorescent Pseudomonas spp. from two different agricultural soils by using three different growth media. CLP production was observed in a large proportion of the strains (approximately 60%) inhabiting the sandy soil, compared to a low proportion (approximately 6%) in the loamy soil. Chemical structure analysis revealed that all CLPs could be clustered into two major groups, each consisting of four subgroups. The two major groups varied primarily in the number of amino acids in the cyclic peptide moiety, while each of the subgroups could be differentiated by substitutions of specific amino acids in the peptide moiety. Production of specific CLPs could be affiliated with Pseudomonas fluorescens strain groups belonging to biotype I, V, or VI. In vitro analysis using both purified CLPs and whole-cell P. fluorescens preparations demonstrated that all CLPs exhibited strong biosurfactant properties and that some also had antibiotic properties towards root-pathogenic microfungi. The CLP-producing P. fluorescens strains provide a useful resource for selection of biological control agents, whether a single strain or a consortium of strains was used to maximize the synergistic effect of multiple antagonistic traits in the inoculum.

Cloning, characterization and expression of a bifunctional fructose-6-phosphate, 2-kinase/fructose-2,6-bisphosphatase from potato.

We have isolated cDNA clones encoding the regulatory enzyme fructose-6-phosphate,2-kinase/fructose-2,6-bisphosphatase from a potato (Solanum tuberosum) leaf cDNA library. All clones represented transcripts of the same gene (F2KP1). Functionality of the encoded protein was verified by expression of the active enzyme in Escherichia coli. The expressed enzyme had both kinase activity which forms fructose-2,6-bisphosphate from fructose-6-phosphate and ATP, and phosphatase activity which degrade fructose-2,6-bisphosphate. The recombinant potato enzyme was radiolabelled by [2-32P]fructose-2,6-bisphosphate verifying conservation of the phosphatase catalytic mechanism which involves a phospho-protein intermediate. The deduced amino acid sequence corresponding to the catalytic core for F2KPI is homologous to the fructose-6-phosphate, 2-kinase/fructose-2,6-bisphosphatase isolated from animals and yeast, with conservation of amino acids involved in substrate binding and catalytic mechanisms. The sequence for F2KP1 also includes a 102 amino acids long NH2-terminal with no homology to any previously identified enzymes. This NH2 terminal may be even longer since an upstream stop codon has not yet been identified. Northern blot analysis of potato showed that the F2KP1 transcript is present in several tissues including source leaves, sink leaves and flowers, whereas the transcripts were not detectable in developing tubers. Southern blot analysis of Solanum phureja suggest there to be only one copy of the gene.