Enzymatic and molecular characterization of an acidic and thermostable chitinase 1 from Streptomyces thermodiastaticus HF 3-3

The Journal of General and Applied Microbiology

Hidehisa Fujiki¹  Wasana Suyotha²  Junji Hayashi³  Keitaro Take⁴ 

[1] Biotechnology for Bioresource Utilization Laboratory, Department of Industrial Biotechnology, Faculty of Agro-industry, Prince of Songkla University;Department of Applied Chemistry, Faculty of Life Sciences, Ritsumeikan University;Department of Biochemical Engineering, Graduate School of Sciences and Engineering, Yamagata University;Department of Biotechnology, Faculty of Life Sciences, Ritsumeikan University

关键词: acidic;    chitinase;    Streptomyces thermodiastaticus;    thermal stability;   

DOI  :  10.2323/jgam.2017.12.002

学科分类：微生物学和免疫学

来源: Applied Microbiology, Molecular and Cellulrar Biosciences Research Foundation

【 摘 要 】

Chitinase 1 (Chi1) is an acidic and thermostable hydrolytic enzyme capable of the breakdown of chitin, a resilient biopolymer that is the primary building block of fungi cell walls and marine exoskeletons. In this study, Chi1 was purified from the bacterium Streptomyces thermodiastaticus HF 3-3, and its properties were carefully characterized. The molecular mass of Chi1 was estimated to be approximately 46 kDa and, through sequencing, its N-terminal amino acid sequence was identified as ADSGKVKL. Although the optimal operating temperature and pH for Chi1 were determined to be 65°C and pH 5.5, respectively, the purified enzyme was stable over wide pH (1.5–9) and temperature ranges. Moreover, Chi1 retained 87% of its activity in the presence of 15% NaCl. While Chi1 activity was inhibited by Ag+ and Mn2+, other chemicals tested had no significant effect on its enzymatic activity. The Km and Vmax values of Chi1 for the substrate colloidal chitin were 1.23 ± 0.7 mg/mL and 6.33 ± 1.0 U/mg, respectively. Thin-layer chromatography analysis of the enzymatic reaction end products mainly detected diacetylchitobiose. We also cloned the Chi1 gene and purified the recombinant protein; the properties of the recombinant enzyme were nearly identical to those of the native enzyme. Therefore, Chi1 purified from S. thermodiastaticus HF 3-3 is unique, as it is highly stable under broad range of pH values, temperatures, and chemical exposures. Combined, these properties make this enzyme attractive for use in the industrial bioconversion of chitin.

【 授权许可】

Unknown