Showing papers by "Alan Collmer published in 1986"
TL;DR: Although there was considerable intraspecific heterogeneity, all strains produced at least four isozymes of pectate lyase, which could be divided into three groups: basic (pI, ca. 9.0 to 10.0), slightly basic ( pI, 7.5 to 8.5), and acidic (p I, 4.8 to 5.0).
Abstract: Erwinia spp that cause soft-rot diseases in plants produce a variety of extracellular pectic enzymes To assess the correlation between patterns of pectic enzyme production and taxonomic classification, we compared the enzymes from representative strains Supernatants obtained from polygalacturonate-grown cultures of nine strains of Erwinia chrysanthemi, three strains of E carotovora subsp carotovora, and three strains of E carotovora subsp atroseptica were concentrated and subjected to ultrathin-layer polyacrylamide gel isoelectric focusing Pectate lyase, polygalacturonase, and exo-poly-alpha-D-galacturonosidase activities were visualized by staining diagnostically buffered pectate-agarose overlays with ruthenium red after incubation of the overlays with the isoelectric focusing gels The isoelectric focusing profiles of pectate lyase and polygalacturonase were nearly identical for strains of E carotovora subsp carotovora and E carotovora subsp atroseptica, showing three pectate lyase isozymes with isoelectric points higher than 87 and a polygalacturonase with pI of ca 102 Isoelectric focusing profiles of the E chrysanthemi pectic enzymes were substantially different Although there was considerable intraspecific heterogeneity, all strains produced at least four isozymes of pectate lyase, which could be divided into three groups: basic (pI, ca 90 to 100), slightly basic (pI, ca 70 to 85), and acidic (pI, ca 40 to 50) Several strains of E chrysanthemi also produced a single form of exo-poly-alpha-D-galacturonosidase (pI, ca 80)(ABSTRACT TRUNCATED AT 250 WORDS)
73 citations
TL;DR: Results indicate that pectate lyase induces a K(+) efflux/H(+) influx response in tobacco similar to but of shorter duration than that induced by P. syringae pv.
Abstract: A purified pectate lyase isozyme derived from Erwinia chrysanthemi induced rapid net K+ efflux and H+ influx in suspension-cultured tobacco cells. Comparable fluxes of other ions (Na+, Cl−) were not observed. The K+ efflux/H+ influx response began within 15 minutes after addition of enzyme to cell suspensions and continued for approximately 1 hour after which cells resumed the net H+ efflux exhibited prior to enzyme treatment. The response was not prolonged by a second enzyme dose 1 hour after the first. The K+/H+ response was characterized by saturation at low enzymic activity (2 × 10−3 units per milliliter), and inhibition by the protonophore, carbonyl cyanide m-chlorophenylhydrazone, and was not associated with membrane leakiness caused by structural cell wall damage. The total K+ loss and H+ uptake induced by enzyme was one-fourth to one-third that induced by Pseudomonas syringae pv. pisi and did not reduce cell viability. These results indicate that pectate lyase induces a K+ efflux/H+ influx response in tobacco similar to but of shorter duration than that induced by P. syringae pv. pisi during the hypersensitive response. Pectate lyase or other cell wall degrading enzymes may therefore influence the induction of hypersensitivity.
24 citations
TL;DR: The genes for two of several pectate lyase isozymes produced by the phytopathogenic enterobacterium Erwinia chrysanthemi 1237 were subcloned and compared by DNA-DNA hybridization, and data indicate that the genes are duplicates and that the proteins are isofunctional.
Abstract: The genes for two of several pectate lyase isozymes produced by the phytopathogenic enterobacterium Erwinia chrysanthemi 1237 were subcloned and compared by DNA-DNA hybridization, and the encoded proteins were analyzed. The borders of the genes were located on a restriction map by incremental exonuclease III deletions. DNA-DNA hybridization studies revealed a low percentage of mismatch (7 to 17%) between pelB and pelC. No homology was detected between pelC and other regions of the E. chrysanthemi 1237 chromosome, in which three other isozyme genes apparently reside. The pectate lyase isozymes were readily purified by chromatofocusing or granulated-gel bed isoelectric focusing from the periplasmic shock fluids of Escherichia coli subclones. The molecular weights of PLb and PLc were 30,000 and 33,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Their isoelectric points were 7.6 and 8.1, respectively, as determined by equilibrium isoelectric focusing in ultrathin polyacrylamide gels. The Km values for PLb and PLc were 0.20 and 0.32 mg/ml, respectively, with polygalacturonate as a substrate. Thin-layer chromatography of reaction products and viscometric assays revealed little difference between the two isozymes. All our data indicate that the genes are duplicates and that the proteins are isofunctional.
23 citations
TL;DR: Heat-stable products released by pectate lyase from isolated tobacco cell walls were similarly able to suppress the hypersensitive response in tobacco leaves, suggesting that the protection phenomenon may be mediated by galacturonic acid-containing oligosaccharides.
21 citations
01 Jan 1986
TL;DR: Experiments with pectate lyases purified from Erwinia cultures have demonstrated that these enzymes can act alone in causing the maceration and host cell killing characteristic of bacterial soft rots.
Abstract: Pectic enzymes have been implicated as pathogenicity factors in a broad range of plant diseases caused by necrotrophic pathogens. The role of pectic enzymes in the soft rots caused by highly pectolytic Erwinia spp. has received particular attention. Experiments with pectate lyases purified from Erwinia cultures have demonstrated that these enzymes can act alone in causing the maceration and host cell killing characteristic of bacterial soft rots [1,2,3,4]. The erwinias are now amenable to a wide array of molecular biological techniques which facilitate exploration of the regulation, export, and role of pectic enzymes in pathogenesis.
7 citations