[Cloning and expression of the gene for thermostable beta-galactosidase from Thermoanaerobacter ethanolicus in Escherichia coli: purification and properties of the product].
An anaerobic thermophilic bacterium Thermoanaerobacter ethanolicus 39E (Clostridium thermohydrosulfuricum 39E) gene library was constructed in E. coli. Recombinant plasmid (pUT50) containing the thermostable beta-galactosidase was isolated by direct selection of clones for enzyme activity using 5-bromo-4-chloro-3-indolyl-D-galactopyranoside (X-gal) and mapping procedures were carried out. The beta-galactosidase was purified from cell extracts of E. coli. Physicochemical characteristics of the recombinant beta-galactosidase were determined. The enzyme has two optimum pH values: 5.3 and 6.0, the temperature optimum is 75-80 degrees C. The molecular weight of beta-galactosidase was determined by PAG electrophoresis: about 83 kDa.
Fokina
N A
NA
Velikodvorskaia
G A
GA
D001421:Bacteria, Anaerobic / Q000201:enzymology*; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D016988:Gram-Positive Asporogenous Rods, Irregular / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D001616:beta-Galactosidase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D011994:Recombinant Proteins; D001616:beta-Galactosidase