A New High-alkaline and High-molecular-weight Pectate Lyase from a Bacillus Isolate : Enzymatic Properties and Cloning of the Gene for the Enzyme

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A pectate lyase (Pel; pectate transeliminase : EC4.2.2.2.), designated Pel-15H, was found in an alkaline culture of Bacillus sp. strain KSM-P15 and purified to homogeneity by sequential column chromatographies. The molecular weight of the enzyme determined by SDS-polyacrylamide gel electrophoresis was approximately 70,000 and the pI was around pH 4.6. Pel-15H randomly trans-eliminated polygalacturonate in the presence of Ca^<2+> ions, and the maximum activity was observed at pH 11.5 and at 55℃ in glycine-NaOH buffer. The gene for Pel-15H was cloned and sequenced, and the structural gene contained a 2,031-bp open reading frame that encoded 677 amino acids including a possible 28-amino-acid signal sequence. The mature enzyme (649 amino acids, molecular weight 69,550) showed very low similarity to Pels from Bacillus with 12.7-18.2% identity. Interestingly, part of the amino acid sequence of Pel-15H had fairly high similarity only to an N-terminal half of PelL and a C-terminal half of PelX from Erwinia chrysanthemi 3937,and a C-terminal half of PelX from E.chrysanthemi EC16 (approximately 35% identity for all).
社団法人日本農芸化学会の論文
2000-06-23