title: Mutations to alter Aspergillus awamori glucoamylase selectivity. II. Mutation of residues 119 and 121.
abstract: Mutations Ser119-->Glu, Ser119-->Gly, Ser119-->Trp, Gly121-->Ala and Gly121-->Ala/Ser411-->Gly were constructed in glucoamylase to change substrate specificity. Mutation Ser411-->Gly was already known to decrease glucoamylase selectivity toward isomaltose formation and to increase peak glucose yield. All mutated glucoamylases had slightly lower specific activities on maltose than on wild-type glucoamylase. Ser119-->Glu, Ser119-->Gly and Ser119-->Trp glucoamylases were about as active on isomaltose and DP 4-7 maltooligosaccharides as wild-type glucoamylase. Gly121-->Ala and Gly121-->Ala/Ser411-->Gly glucoamylases were less active. At 55 degrees C Ser119-->Glu, wild-type, Ser119-->Trp, Ser119-->Gly, Gly121-->Ala and Gly121-->Ala/Ser411-->Gly glucoamylases had progressively higher peak glucose yields, generally in the opposite order to their activities. There was also an inverse correlation between peak glucose yield and ratio of initial rate of isomaltose production from glucose condensation to that of glucose production from maltodextrin hydrolysis. The effect of mutations Gly121-->Ala and Ser411-->Gly was not additive in predicting the effect of the double mutation on the ratio or on peak glucose yield.
authors.affiliation: Department of Food Science and Human Nutrition, Iowa State University, Ames 50011, USA.
authors.forename: T Y
authors.initials: TY
authors.lastname: Fang
authors.forename: R B
authors.initials: RB
authors.lastname: Honzatko
authors.forename: P J
authors.initials: PJ
authors.lastname: Reilly
authors.forename: C
authors.initials: C
authors.lastname: Ford
mesh_terms: D001230:Aspergillus / Q000201:enzymology*; D002236:Carbohydrate Conformation; D005087:Glucan 1,4-alpha-Glucosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D005998:Glycine; D006868:Hydrolysis; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D011134:Polysaccharides / Q000378:metabolism; D012694:Serine; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D011134:Polysaccharides; D012694:Serine; C008315:maltodextrin; D005087:Glucan 1,4-alpha-Glucosidase; D005947:Glucose; D005998:Glycine