title: Improvement in the catalytic activity of β-agarase AgaA from Zobellia galactanivorans by site-directed mutagenesis.
abstract: In this study, site-directed mutagenesis was performed on the β-agarase AgaA gene from Zobellia galactanivorans to improve its catalytic activity and thermostability. The activities of three mutant enzymes, S63K, C253I, and S63K-C253I, were 126% (1,757.78 U/mg), 2.4% (33.47 U/mg), and 0.57% (8.01 U/mg), respectively, relative to the wildtype beta-agarase AgaA (1,392.61 U/mg) at 40°C. The stability of the mutant S63K enzyme was 125% of the wild-type up to 45°C, where agar is in a sol state. The mutant S63K enzyme produced 166%, 257%, and 220% more neoagarohexaose, and 230%, 427%, and 350% more neoagarotetraose than the wild-type in sol, gel, and nonmelted powder agar, respectively, at 45°C over 24 h. The mutant S63K enzyme produced 50% more neoagarooligosaccharides from agar than the wild-type beta-agarase AgaA from agarose under the same conditions. Thus, mutant S63K β-agarase AgaA may be useful for the production of functional neoagarooligosaccharides.
authors: Department of Pharmaceutical Engineering, College of Medical Life Sciences, Silla University, Busan 617-736, Korea.
authors: Seungwoo
authors: S
authors: Lee
authors: Dong-Geun
authors: DG
authors: Lee
authors: Min-Kyung
authors: MK
authors: Jang
authors: Myong-Je
authors: MJ
authors: Jeon
authors: Hye-Ji
authors: HJ
authors: Jang
authors: Sang-Hyeon
authors: SH
authors: Lee
mesh_terms: D000595:Amino Acid Sequence; D004795:Enzyme Stability; D041965:Flavobacteriaceae / Q000201:enzymology*; D005697:Galactosides / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism; D016415:Sequence Alignment; D013696:Temperature; D013997:Time Factors
chemical_list: D005697:Galactosides; D050505:Mutant Proteins; D009844:Oligosaccharides; D006026:Glycoside Hydrolases; C019624:agarase