title: Cloning and sequencing of a gene encoding the 69-kDa extracellular chitinase of Janthinobacterium lividum.
abstract: Janthinobacterium lividum secretes a major 56-kDa chitinase and a minor 69-kDa chitinase. A chitinase gene was defined on a 3-kb fragment of clone pRKT10, by virtue of fluorescent colonies in the presence of 4-methylumbelliferyl-beta-D-N,N',N"-chitotrioside. Nucleotide sequencing revealed an 1998-bp open reading frame with the potential to encode a 69,716-Da protein with amino acid sequences similar to those in other chitinases, suggesting it encodes the minor chitinase (Chi69). Chitinase activity of Escherichia coli (pRKT10) lysates was detected mainly in the periplasmic fraction and immunoblotting detected a 70-kDa protein in this fraction. Chi69 has an N-terminal secretory leader peptide preceding two probable chitin-binding domains and a catalytic domain. These functional domains are separated by linker regions of proline-threonine repeats. Amino acid sequencing of cyanogen bromide cleavage-derived peptides from the major 56-kDa chitinase suggested that Chi69 may be a precursor of Chi56. In addition, an N-terminally truncated version of Chi69 retained chitinase activity as expected if in vivo processing of Chi69 generates Chi56.
authors.affiliation: Gene Transfer and Expression Group, Horticulture and Food Research Institute of New Zealand Ltd., Auckland.
authors.forename: A P
authors.initials: AP
authors.lastname: Gleave
authors.forename: R K
authors.initials: RK
authors.lastname: Taylor
authors.forename: B A
authors.initials: BA
authors.lastname: Morris
authors.forename: D R
authors.initials: DR
authors.lastname: Greenwood
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002686:Chitin / Q000378:metabolism; D002688:Chitinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics*; D006088:Gram-Negative Aerobic Bacteria / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D017421:Sequence Analysis; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
chemical_list: D002686:Chitin; D002688:Chitinases