Analysis of bgl operon structure and characterization of beta-glucosidase from Pectobacterium carotovorum subsp. carotovorum LY34.
A putative bgl operon of Pectobacterium carotovorum subsp. carotovorum LY34 (Pcc LY34) was isolated. Sequence analysis of the 5,557 bp cloned DNA fragment (accession no. AY542524) showed three open reading frames (bglT, bglP, and bglB) predicted to encode 287, 633, and 468 amino acid proteins respectively. BglT and BglP ORFs show high similarity to that of the Pectobacterium chrysanthemi ArbG antiterminator and ArbF permease respectively. Also, the latter contains most residues important for phosphotransferase activity. The amino acid sequence of BglB showed high similarity to various beta-glucosidases and is a member of glycosyl hydrolase family 1. The purified BglB enzyme hydrolyzed salicin, arbutin, pNPG, and MUG. The molecular weight of the enzyme was estimated to be 53,000 Da by SDS-PAGE. The purified beta-glucosidase exhibited maximal activity at pH 7.0 and 40 degrees C, and its activity was enhanced in the presence of Mg(2+). Two glutamate residues (Glu(173) and Glu(362)) were found to be essential for enzyme activity.
An
Chang Long
CL
Division of Applied Life Science, Gyeongsang National University, Chinju, Korea.
Lim
Woo Jin
WJ
Hong
Su Young
SY
Kim
Eun Ju
EJ
Shin
Eun Chule
EC
Kim
Min Keun
MK
Lee
Jong Reoul
JR
Park
Sang Ryeol
SR
Woo
Jong Gyu
JG
Lim
Yong Pyo
YP
Yun
Han Dae
HD
D001483:Base Sequence; D003001:Cloning, Molecular; D003360:Cosmids; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D015723:Gene Library; D026901:Membrane Transport Proteins / Q000378:metabolism; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009876:Operon / Q000235:genetics*; D044043:Pectobacterium / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development; D011995:Recombination, Genetic; D013696:Temperature; D001617:beta-Glucosidase / Q000235:genetics*
D004269:DNA, Bacterial; D026901:Membrane Transport Proteins; D008670:Metals; D001617:beta-Glucosidase