Purification and partial characterization of a mitomycin C-induced pectin lyase of Erwinia chrysanthemi strain EC183

Abstract

Pectin lyase, produced by Erwinia chrysanthemi strain EC183 following mitomycin C treatment, was purified to near homogeneity from the culture lysate. The purification procedure involved ultrafiltration, ammonium sulphate fractionation and ion exchange chromatography on DEAE-BioGel A and CM-sepharose. The molecular weight and pI of the pectin lyase were estimated to be 34 500 and 9·45, respectively. The enzyme was most active on Link (98% esterified) pectin and least active on nonesterified pectic acid. Pectin lyase cleaved Link pectin in an endo manner, with optimum pH of 8·3 and optimum temperature of 33 †C. The K_m and V_max of the enzyme with Link pectin as the substrate were estimated to be 7·3±2·0 mg ml⁻¹ and 47·6±10·7 A₂₃₅ min⁻¹, respectively. Antipectin lyase antibody reacted with pectin lyases from 11 of 12 E. chrysanthemi strains and all four Erwinia carotovora subsp. atroseptica strains tested but not with enzymatic preparations from E. carotovora subsp. carotovora, Erwinia milletiae, Erwinia rhapontici or Aspergillus sojae. These data indicate that pectin lyase of E. carotovora subsp. carotovora is immunologically distinct from that produced by E. chrysanthemi.