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  • 1
    ISSN: 1432-2048
    Keywords: Cell wall ; Extensin ; Extensin peroxidase ; Hydroxyproline-rich glycoprotein ; Lycopersicon (cell culture)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Extensin, a hydroxyproline-rich glycoprotein comprising substantial amounts of β-l-arabinose-hydroxyproline glycosidic linkages is believed to be insolubilized in the cell wall during host-pathogen interaction by a peroxidase/hydroperoxide-mediated cross-linking process. Both extensin precursor and extensin peroxidase were ionically eluted from intact water-washed tomato (hybrid) of Lycopersicon esculentum Mill. and L. peruvianum L. (Mill.) cells in suspension cultures and purified to homogeneity by a rapid and simple procedure under mild and non-destructive experimental conditions. The molecular weight of native extensin precursor was estimated to be greater than 240–300 kDa by Superose-12 gel-filtration chromatography. Extensin monomers have previously been designated a molecular weight of approximately 80 kDa. Our results indicate that salt-eluted extensin precursor is not monomeric. Agarose-gel electrophoresis, Superose-12-gel-filtration, extensin-peroxidase-catalysed cross-linking, Mono-S ion-exchange fast protein liquid chromatography (FPLC), and peptide-sequencing data confirmed the homogeneity of the extensin preparation. Evidence that the purified protein was extensin is attributed to the presence of the putative sequence motif — Ser (Hyp)4 — within the N-terminal end of the protein. Treatment of extensin with trifluoroacetic acid demonstrated that arabinose was the principal carbohydrate. The amino-acid composition of the purified extensin was similar to those reported in the literature. The cross-linking of extensin in vitro upon incubation with extensin peroxidase and exogenous H2O2 was characteristic of other reported extensins. Furthermore, Mono-S ion-exchange FPLC of native extensin precursor resolved it into two isoforms, A (90%) and B (10%). The amino-acid compositions of extensin A and extensin B were found to be similar to each other and both extensins were cross-linked in vitro by extensin peroxidase.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-2048
    Keywords: Chalcone synthase ; Colletotrichum ; Elicitor ; Phaseolus ; Phenylalanine ammonia-lyase ; Phytoalexin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In wounded cotyledons ofPhaseolus vulgaris L. the accumulation of the 5-hydroxy isoflavonoids kievitone and 2′-hydroxygenistein precedes the major increases in the levels of the 5-deoxy compounds phaseollin and coumestrol. Increased phytoalexin levels are preceded by transient increases in the extractable activities of L-phenylalanine ammonia-lyase (EC 4.3.1.5.), chalcone synthase and chalcone isomerase (EC 5.5.1.6.). Accumulation of phytoalexins, above wounded control levels, is observed following treatment of excised cotyledons or hypocotyls with crude or fractionated elicitor preparations heat-released from the cell walls ofColletotrichum lindemuthianum. Chalcone synthase levels are also induced in cotyledons, although crude elicitor and all fractions suppress L-phenylalanine ammonia-lyase activity in both tissues. Kievitone is the major phytoalexin induced in cotyledons, whereas in hypocotyls phaseollin predominates. Patterns of phytoalexin accumulation have been studied in response to varying concentrations of the crude and fractionated elicitor; 5-hydroxy isoflavonoid accumulation is highly dependent upon elicitor concentration, the dose-response curves for kievitone accumulation showing maxima at around 1 μg glucose equivalents per cotyledon, minima at 2–3 μg equivalents and increasing induction at higher concentrations. Similar patterns are observed for L-phenylalanine ammonia-lyase and chalcone synthase levels, although the overall extent of these changes is masked by the high wound response. Accumulation of 5-deoxy isoflavonoids above control levels requires high elicitor concentrations; no experimental conditions were found under which phaseollin accumulated to higher levels than kievitone in cotyledons during the first 48 h after elicitation.
    Type of Medium: Electronic Resource
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