title: Enhancement of the thermostability of the maltogenic amylase MAUS149 by Gly312Ala and Lys436Arg substitutions.
abstract: Based on sequence alignments and homology modeling, Gly 312 and Lys 436 of the maltogenic amylase from Bacillus sp. US149 (MAUS149) were selected as targets for site-directed mutagenesis to improve the thermostability of the enzyme. Variants of MAUS149 with amino acid substitutions G312A, K436R and G312A-K436R had substrate specificities, kinetic parameters and pH optima similar to those of the wild-type enzyme; however, the enzymes with substitutions K436R and G312A-K436R, had an optimal temperature of 45 °C instead of the 40 °C for the wild-type enzyme. The half-life time at 55 °C increased from 15 to 25 min for the double mutant. Molecular modeling suggests that the increase in thermostability was due to new hydrophobic interactions and the formation of a salt bridge and hydrogen bond in the G312A and K436R variants, respectively. The double mutant could be a potential candidate for application in the bread industry.
authors.affiliation: Laboratoire de Microorganismes et de Biomolécules, Centre de Biotechnologie de Sfax, BP 1177, Sfax 3018, Tunisia.
authors.forename: Sameh
authors.initials: S
authors.lastname: Ben Mabrouk
authors.forename: Nushin
authors.initials: N
authors.lastname: Aghajari
authors.forename: Mamdouh
authors.initials: M
authors.lastname: Ben Ali
authors.forename: Ezzedine
authors.initials: E
authors.lastname: Ben Messaoud
authors.forename: Michel
authors.initials: M
authors.lastname: Juy
authors.forename: Richard
authors.initials: R
authors.lastname: Haser
authors.forename: Samir
authors.initials: S
authors.lastname: Bejar
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics*; D000596:Amino Acids / Q000235:genetics*; D001407:Bacillus / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000379:methods*; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D009154:Mutation / Q000235:genetics; D016415:Sequence Alignment; D013696:Temperature
chemical_list: D000596:Amino Acids; D050505:Mutant Proteins; D006026:Glycoside Hydrolases; C120909:glucan 1,4-alpha-maltohydrolase