title: Cloning and characterization of katA, encoding the major monofunctional catalase from Xanthomonas campestris pv. phaseoli and characterization of the encoded catalase KatA.
abstract: The first cloning and characterization of the gene katA, encoding the major catalase (KatA), from Xanthomonas is reported. A reverse genetic approach using a synthesized katA-specific DNA probe to screen a X. campestris pv. phaseoli genomic library was employed. A positively hybridizing clone designated pKat29 that contained a full-length katA was isolated. Analysis of the nucleotide sequence revealed an open reading frame of 1,521 bp encoding a 507-amino acid protein with a theoretical molecular mass of 56 kDa. The deduced amino acid sequence of KatA revealed 84% and 78% identity to CatF of Pseudomonas syringae and KatB of P. aeruginosa, respectively. Phylogenetic analysis places Xanthomonas katA in the clade I group of bacterial catalases. Unexpectedly, expression of katA in a heterologous Escherichia coli host resulted in a temperature-sensitive expression. The KatA enzyme was purified from an overproducing mutant of X. campestris and was characterized. It has apparent K(m) and V(max) values of 75 m M [H(2)O(2)] and 2.55 x 10(5) micromol H(2)O(2) micromol heme(-1) s(-1), respectively. The enzyme is highly sensitive to 3-amino-1,2,4-triazole and NaN(3), has a narrower optimal pH range than other catalases, and is more sensitive to heat inactivation.
authors.affiliation: Department of Biotechnology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Nopmanee
authors.initials: N
authors.lastname: Chauvatcharin
authors.forename: Paiboon
authors.initials: P
authors.lastname: Vattanaviboon
authors.forename: Jack
authors.initials: J
authors.lastname: Switala
authors.forename: Peter C
authors.initials: PC
authors.lastname: Loewen
authors.forename: Skorn
authors.initials: S
authors.lastname: Mongkolsuk
mesh_terms: D002374:Catalase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis; D006861:Hydrogen Peroxide / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D016959:Xanthomonas campestris / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development
chemical_list: D004269:DNA, Bacterial; D006861:Hydrogen Peroxide; D002374:Catalase