Cloning and expression of the gene encoding Streptomyces coelicolor A3(2) alpha-galactosidase belonging to family 36.
The alpha-galactosidase gene of Streptomyces coelicolor A3(2) was cloned, expressed in Escherichia coli and characterized. It consisted of 1497 nucleotides encoding a protein of 499 amino acids with a predicted molecular weight of 57,385. The observed homology between the deduced amino acid sequences of the enzyme and alpha-galactosidase from Thermus thermophilus was over 40%. The alpha-galactosidase gene was assigned to family 36 of the glycosyl hydrolases. The enzyme purified from recombinant E. coli showed optimal activity at 40 degrees C and pH 7. The enzyme hydrolyzed p-nitrophenyl-alpha-D -galactopyroside, raffinose, stachyose but not melibiose and galactomanno-oligosaccharides, indicating that this enzyme recognizes not only the galactose moiety but also other substrates.
Graduated school of Life and Environmental Science, University of Tsukuba, Ibaraki, 305-0006, Japan.
Kenji
K
Kondoh
Kohei
K
Morisaki
Wook-Dong
WD
Kim
Gwi-Gun
GG
Park
Satoshi
S
Kaneko
Hideyuki
H
Kobayashi
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D005690:Galactose / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008553:Melibiose / Q000737:chemistry; D008969:Molecular Sequence Data; D009699:N-Glycosyl Hydrolases / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D048372:Streptomyces coelicolor / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D013696:Temperature; D013824:Thermus; D016963:Thermus thermophilus; D000519:alpha-Galactosidase / Q000096:biosynthesis* / Q000235:genetics*
D009844:Oligosaccharides; D008553:Melibiose; D000519:alpha-Galactosidase; D009699:N-Glycosyl Hydrolases; D005690:Galactose