title: Improvement of thermostability and halostability of β-1,3-1,4-glucanase by substituting hydrophobic residue for Lys48.
abstract: The aim of this study was to improve the stability of β-1,3-1,4-glucanase by substituting hydrophobic residue for specific amino acid. The results indicated that the catalytic efficiency, thermostability and halostability were enhanced simultaneously by replacement of Lys48 with Ala (K48A) or Leu (K48L). Comparison of kinetic parameters revealed that catalytic efficiency of mutants is enhanced as a result of the increase in substrate affinity. A great improvement in thermostability and halostability was observed. The half-lives of mutants significantly increased (up to ∼7-fold) at 60-70°C. Moreover, relative enzymatic activities of mutants were observed more than 80% even in the presence of 30% NaCl, and half-lives were increased to 3-fold that of wild-type. Based on above results, when applied to ionic liquid, mutants were more active and stable compared to wild-type. These were the results from improvement of protein functions by the substitution of hydrophobic single residue in adjacent with forming carbohydrate binding cavity. Therefore, this report could be helpful for improvement of the enzyme property and for biotechnological application as well.
authors.affiliation: Department of Biotechnology, Pukyong National University, Busan, 608-737, Republic of Korea.
authors.forename: Jong Min
authors.initials: JM
authors.lastname: Lee
authors.affiliation: Department of Biotechnology, Pukyong National University, Busan, 608-737, Republic of Korea.
authors.forename: Soo Young
authors.initials: SY
authors.lastname: Moon
authors.affiliation: CBR Defense Research Institute, The Armed Forces CBR Defense Command, Seoul, 137-180, Republic of Korea.
authors.forename: Yu-Ri
authors.initials: YR
authors.lastname: Kim
authors.affiliation: Aquafeed Research Center, NIFS, Pohang, 791-923, Republic of Korea.
authors.forename: Kang Woong
authors.initials: KW
authors.lastname: Kim
authors.affiliation: Aquafeed Research Center, NIFS, Pohang, 791-923, Republic of Korea.
authors.forename: Bong-Joo
authors.initials: BJ
authors.lastname: Lee
authors.affiliation: Department of Biotechnology, Pukyong National University, Busan, 608-737, Republic of Korea. Electronic address: iskong@pknu.ac.kr.
authors.forename: In-Soo
authors.initials: IS
authors.lastname: Kong
mesh_terms: D000409:Alanine / Q000737:chemistry*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001407:Bacillus / Q000737:chemistry* / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D055162:Biocatalysis; D044644:Endo-1,3(4)-beta-Glucanase / Q000737:chemistry* / Q000235:genetics; D004795:Enzyme Stability; D006358:Hot Temperature; D057927:Hydrophobic and Hydrophilic Interactions; D052578:Ionic Liquids / Q000737:chemistry; D007700:Kinetics; D007930:Leucine / Q000737:chemistry*; D008239:Lysine / Q000737:chemistry*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D016415:Sequence Alignment; D012965:Sodium Chloride / Q000737:chemistry; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D052578:Ionic Liquids; D012965:Sodium Chloride; D044644:Endo-1,3(4)-beta-Glucanase; D007930:Leucine; D008239:Lysine; D000409:Alanine
keywords: Halostability; Thermostability; β-1,3-1,4-Glucanase