Purification and characterization of pectic enzymes from two races of Fusarium oxysporum f. sp. ciceri differing in virulence to chickpea (Cicer arietinum L.)

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Abstract

Polygalacturonases (PG) and pectate lyases (PL) produced in vitro by races 0 and 5 of Fusarium oxysporum f. sp. ciceri have been purified by gel filtration followed by anion-exchange chromatography and, in some instances, chromatofocusing. Race 0, the least virulent race, produced three PG forms, designated PG I0, PG II0 and PG III0, and one PL form, designated PL0, whereas race 5, the more virulent race, produced only one PG form, designated PG5, and two PL forms, designated PL I5 and PL II5. The molecular weights of the enzymes were estimated to be 44000 for PG I0 and PG II0 76000 for PG III0 and PG5, 25000 for PL0, and 37000 for PL I5 and PL II5. By their mode of action, PG I0 and PG II0 were endo-enzymes, PG III0 and PG5 were exo-enzymes, and the three lyases were endo-enzymes showing, in contrast to the endo-PGs, very low or null activity on oligogalacturonides shorter than pentamers. With polygalacturonic acid as substrate, the values of pH optimum were 4·5 for all four PG forms, 9·5 for PL0, and 10·5 for PL I5 and PL II5; the PL forms showed an absolute Ca2+ requirement for activity. PG I0, PL0 and PL I5 were cationic proteins with alkaline isoelectric points, whereas PG II0, PG III0, PG5, and PL II5 were anionic proteins with acidic isoelectric points. All the purified pectic enzymes were able to degrade chickpea cell walls although the degradative activity of the exo-PG5 and the anionic endo-PL II5 was notably lower than that of the remaining PG and PL forms. Degradability of wall preparations without ionically bound proteins was not related to host resistance or susceptibility. In contrast, walls containing ionically bound proteins from the chickpea susceptible cv. PV-24 were more extensively degraded by the various PG and PL forms than were similar preparations from the resistant cv. WR-315.

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