Cloning and expression of the gene encoding Streptomyces coelicolor A3(2) α-galactosidase belonging to family 36

Abstract

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.

Solid-state fermentation-supported enhanced production of α-galactosidase by a deoxyglucose-resistant mutant of Aspergillus niger and thermostabilization of the production process

“…The product yield and specific productivity of a-Gal supported by A. niger mutant derivative in the presence of corn steep liquor and glucose (2 g/flask) in SSF are several-fold higher than the reported values by other workers on Aspergillus spp. (den Herder et al 1992;Fridjonsson et al 1999;Kondoh et al 2005). This may have occurred due to the positive regulation of genes involved in regulation of substrate and nitrogen source consumption (Bohm and Boos 2004) in the culture medium.…”

Cloning and Functional Expression of an α-Galactosidase fromYersinia pestis biovar Microtusstr. 91001

“…2C), close to that of the AgaA from Carnobacterium piscicola BA (32-37 C) 10) but lower than most GH-36 -galactosidases from bacteria. 3) After incubation at 50 C for 2, 5, and 10 min, about 63, 10, and 3% of activity was retained respectively (Fig. 2D).…”

“…This suggests that the Cys residue is located at the catalytic site, since these ions have been reported to attack Cys residues at active sites in GH-36 -galactosidases. 3,9) Kinetic studies were performed with pNP--Gal as substrate at 37 C for 2 min. The Michaelis constant (K m ) and catalytic constant (k cat ) were 0.51 mM and 3,279/s.…”

“…91001; characterization -Galactosidase (-D-galactoside galactohydrolase; EC 3.2.1.22) is widely distributed in nature 1) and has been utilized in many industries, and in medical treatment and scientific research. [2][3][4][5] It is also involved in cell-wall modification during tissue development. 6) Based on amino acid sequence similarities, -galactosidases have been classified into glycoside hydrolase (GH) families 4, 27, 36, and 57 in the CAZy database (http://www.cazy.org/fam/acc GH.html).…”

Gene cloning, expression and characterization of an α-galactosidase from Pedobacter nyackensis MJ11 CGMCC 2503 with potential as an aquatic feed additive

“…a-Galactosidase (a-D-galactoside galactohydrolase; EC 3.2.1.22) has been isolated from various sources including microorganisms (Ademark et al 2001;Kondoh et al 2005), plants (Gao and Schaffer 1999;Li et al 2007) and mammals (Bishop et al 1986). Those from microorganisms, such as fungi (Mi et al 2007), bacteria (Silvestroni et al 2002) and archaea (Brouns et al 2006), have received significant attention due to their high yield.…”