TY - JOUR
T1 - Three alfa-galactosidase genes of Trichoderma reesei cloned by expression in yeast
AU - Margolles-Clark, Emilio
AU - Tenkanen, Maija
AU - Luonteri, Elina
AU - Penttilä, Merja
N1 - LIS: A1
CA2: 1503
CA2: 1501
CA: BEL
Project code: BEL4103
PY - 1996
Y1 - 1996
N2 - Three alpha-galactosidase genes, agl1, agl2 and agl3, were isolated from a cDNA expression library of Trichoderma reesei RutC-30 constructed in the yeast Saccharomyces cerevisiae by screening the library on plates containing the substrate 5-bromo-4-chloro-3-indolyl-alpha-D-galactopyranoside. The genes agl1, agl2 and agl3 encode 444, 746 and 624 amino acids, respectively, including the signal sequences. The deduced amino acid sequences of AGLI and AGLIII showed similarity with the alpha-galactosidases of plant, animal, yeast and filamentous fungal origin classified into family 27 of glycosyl hydrolases whereas the deduced amino acid sequence of AGLII showed similarity with the bacterial alpha-galactosidases of family 36. The enzymes produced by yeast were analysed for enzymatic activity against different substrates. AGLI, AGLII and AGLIII were able to hydrolyse the synthetic substrate p-nitrophenyl-alpha-D-galactopyranoside and the small galactose-containing oligosaccharides, melibiose and raffinose. They liberated galactose from polymeric galacto(gluco)mannan with different efficiencies. The action of AGLI towards polymeric substrates was enhanced by the presence of the endo-1,4-beta-mannanase of T. reesei. AGLII and AGLIII showed synergy in galacto(gluco)mannan hydrolysis with the endo-1,4-beta-mannanase of T. reesei and a beta-mannosidase of Aspergillus niger. The calculated molecular mass and the hydrolytic properties of AGLI indicate that it corresponds to the alpha-galactosidase previously purified from T. reesei.
AB - Three alpha-galactosidase genes, agl1, agl2 and agl3, were isolated from a cDNA expression library of Trichoderma reesei RutC-30 constructed in the yeast Saccharomyces cerevisiae by screening the library on plates containing the substrate 5-bromo-4-chloro-3-indolyl-alpha-D-galactopyranoside. The genes agl1, agl2 and agl3 encode 444, 746 and 624 amino acids, respectively, including the signal sequences. The deduced amino acid sequences of AGLI and AGLIII showed similarity with the alpha-galactosidases of plant, animal, yeast and filamentous fungal origin classified into family 27 of glycosyl hydrolases whereas the deduced amino acid sequence of AGLII showed similarity with the bacterial alpha-galactosidases of family 36. The enzymes produced by yeast were analysed for enzymatic activity against different substrates. AGLI, AGLII and AGLIII were able to hydrolyse the synthetic substrate p-nitrophenyl-alpha-D-galactopyranoside and the small galactose-containing oligosaccharides, melibiose and raffinose. They liberated galactose from polymeric galacto(gluco)mannan with different efficiencies. The action of AGLI towards polymeric substrates was enhanced by the presence of the endo-1,4-beta-mannanase of T. reesei. AGLII and AGLIII showed synergy in galacto(gluco)mannan hydrolysis with the endo-1,4-beta-mannanase of T. reesei and a beta-mannosidase of Aspergillus niger. The calculated molecular mass and the hydrolytic properties of AGLI indicate that it corresponds to the alpha-galactosidase previously purified from T. reesei.
U2 - 10.1111/j.1432-1033.1996.0104h.x
DO - 10.1111/j.1432-1033.1996.0104h.x
M3 - Article
SN - 0014-2956
VL - 240
SP - 104
EP - 111
JO - European Journal of Biochemistry
JF - European Journal of Biochemistry
IS - 1
ER -