title: Activation of a casB gene encoding β-glucosidase of Pectobacterium carotovorum subsp. carotovorum LY34.
abstract: Two cas genes were isolated from Pectobacterium carotovorum subsp. carotovorum LY34 (Pcc LY34). Sequence analysis of the 4873 bp cloned DNA fragment (accession number AY866383) revealed two open reading frames (casF and casB) that are predicted to encode 658 and 467 amino acid proteins, respectively. The CasF protein is similar to other PTS enzyme II components. casB encodes β-glucosidase, a member of the glycosyl hydrolase family 1. An inverted repeat sequence was identified in the casB promoter region, and was hypothesized to have a negative effect on casB transcription. Replacement of the casB promoter of Pcc LY34 with the bglB promoter activated the casB gene, consistent with the repeats inhibiting expression of casB. Purified CasB enzyme was estimated to be 53,000 Da by SDS-PAGE, and hydrolyzed salicin, arbutin, pNPG, and MUG. CasB exhibited maximal activity toward pNPG at pH 7.0 and 40 °C, and Mg(2+) is essential for its activity. Two conserved glutamate residues (Glu(177) and Glu(366)) were shown to be important for CasB activity.
authors.affiliation: Gyeongsangnam-do Agricultural Research and Extension Service, Chinju 660-360, Republic of Korea.
authors.forename: Min Keun
authors.initials: MK
authors.lastname: Kim
authors.forename: Chang Long
authors.initials: CL
authors.lastname: An
authors.forename: Tae Ho
authors.initials: TH
authors.lastname: Kang
authors.forename: Jungho
authors.initials: J
authors.lastname: Kim
authors.forename: Hoon
authors.initials: H
authors.lastname: Kim
authors.forename: Han Dae
authors.initials: HD
authors.lastname: Yun
mesh_terms: D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D055029:Inverted Repeat Sequences; D060847:Metabolic Engineering; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D016973:Pectobacterium carotovorum / Q000201:enzymology* / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D013696:Temperature; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D001617:beta-Glucosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D004269:DNA, Bacterial; D001617:beta-Glucosidase