title: Nucleotide and amino-acid sequences of a new-type pectate lyase from an alkaliphilic strain of Bacillus.
abstract: A pectate lyase (pectate transeliminase; EC 4.2.2.2), designated Pel-15E, was purified to homogeneity from a culture broth of alkaliphilic Bacillus sp. strain KSM-P15. The purified enzyme had a molecular mass of approximately 33 kDa, as determined by SDS/PAGE, and a pI of approximately pH 9.2. Pel-15E exhibited optimum activity at pH 10.5 and 50-55 degrees C in glycine/NaOH buffer. Pel-15E had an absolute requirement for Ca2+ ions for manifestation of the enzymatic activity and trans-eliminated poly(galacturonic) acid, most likely by endo-type cleavage. A gene for the enzyme, which was cloned using the shotgun method and sequenced, contained a 960-bp ORF encoding 320 amino acids. The mature enzyme (286 amino acids, 32 085 Da) from the deduced amino-acid sequence showed quite low homology to known Pels from various microorganisms with 16.1-20.4% identity. Furthermore, we were not able to find any conserved regions in the sequence of Pel-15E when aligned with the sequences of other enzymes from the established Pel superfamily. However, Pel-15E had some regions that were homologous to PelA from Azospirillum irakense with 39.8% identity. Based on their amino-acid sequence homology, Pel-15E and PelA appear to belong to a new class of Pel family, although the enzymatic properties of both enzymes were quite different.
authors.affiliation: Tochigi Research Laboratories of Kao Corporation, Tochigi, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Sawada
authors.forename: A
authors.initials: A
authors.lastname: Ogawa
authors.forename: T
authors.initials: T
authors.lastname: Ozawa
authors.forename: N
authors.initials: N
authors.lastname: Sumitomo
authors.forename: Y
authors.initials: Y
authors.lastname: Hatada
authors.forename: T
authors.initials: T
authors.lastname: Kobayashi
authors.forename: S
authors.initials: S
authors.lastname: Ito
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology*; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D008969:Molecular Sequence Data; D011133:Polysaccharide-Lyases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification
chemical_list: D004269:DNA, Bacterial; D011133:Polysaccharide-Lyases; C023911:pectate lyase