Isolation of a beta-galactosidase-encoding gene from Bacillus licheniformis: purification and characterization of the recombinant enzyme expressed in Escherichia coli.
The Bacillus licheniformis beta-galactosidase gene, lacBl, was cloned on a 5.8-kb HindIII fragment into pBR322 and expressed by its own promoter in Escherichia coli. Deletion and complementation analysis showed that the enzyme-encoding region was located on a 4. 1-kb HindIII-ClaI fragment. The transcription region for the lacBl was identified on this fragment with promoter- and terminator-probe plasmids. The deduced sequence of 149 aa of the N-terminal part of lacBl showed aa sequence homology with beta-Gal from B. stearothermophilus, B. circulans, Haloferax alicantei, Clostridium perfringens, Arthrobacter sp.. No significant homology was shared with those found in the lacZ and lacS families. The recombinant beta-galactosidase (LacB1) was purified by FPLC. The molecular mass of the enzyme (80 kDa) and its optimal pH (5.7) and temperature (45 degrees C) were determined.
Trân
L S
LS
Department of Biotechnology and Molecular Genetics, University of Agricultural Sciences, Gödöllõ, Hungary, H-2103, USA.
Szabó
L
L
Fülöp
L
L
Orosz
L
L
Sík
T
T
Holczinger
A
A
D001407:Bacillus / Q000201:enzymology*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification*; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification; D015183:Restriction Mapping; D016415:Sequence Alignment; D013696:Temperature; D001616:beta-Galactosidase / Q000235:genetics / Q000302:isolation & purification*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D001616:beta-Galactosidase