A new high-alkaline and high-molecular-weight pectate lyase from a Bacillus isolate: Enzymatic properties and cloning of the gene for the enzyme

Citation
A. Ogawa et al., A new high-alkaline and high-molecular-weight pectate lyase from a Bacillus isolate: Enzymatic properties and cloning of the gene for the enzyme, BIOS BIOT B, 64(6), 2000, pp. 1133-1141
Citations number
31
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Agricultural Chemistry","Biochemistry & Biophysics
Journal title
BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY
ISSN journal
0916-8451 → ACNP
Volume
64
Issue
6
Year of publication
2000
Pages
1133 - 1141
Database
ISI
SICI code
0916-8451(200006)64:6<1133:ANHAHP>2.0.ZU;2-V
Abstract
A pectate lyase (Pel; pectate transeliminase: EC4.2.2.2.), designated Pel-1 5W, was found in an alkaline culture of Bacillus sp. strain KSM-P15 and pur ified to homogeneity by sequential column chromatographies. The molecular w eight of the enzyme determined by SDS-polyacrylamide gel electrophoresis wa s approximately 70,000 and the pI was around pH 4.6. Pel-15H randomly trans -eliminated polygalacturonate in the presence of Ca2+ ions, and the maximum activity was observed at pH 11.5 and at 55 degrees C in glycine-NaOH buffe r. The gene for Pel-15H was cloned and sequenced, and the structural gene c ontained a 2,031-bp open reading frame that encoded 677 amino acids includi ng a possible 28-amino-acid signal sequence. The mature enzyme (649 amino a cids, molecular weight 69,550) showed very low similarity to Pels from Baci llus with 12.7-18.2% identity, Interestingly, part of the amino acid sequen ce 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-term inal half of PelX from E. chrysanthemi EC16 (approximately 35% identity for all).