Optimization of the fermentation conditions for the mutant strain of β-cyclodextrin glycosyltransferase H167C to produce cyclodextrins

Abstract

The cyclodextrin glycosyltransferase (CGTase) was used to catalyze the conversion of starch into cyclodextrins (CD) in industry. Improving the activity of CGTase to produce more CD with relative low cost is intensely interesting and has drawn wide attention. Amino acid mutation of His167 into Cys significantly enhanced β-CGTase activity; however, optimization of culture conditions for β-CGTase-H167C remains unclear. To determine this, the medium and culture conditions for β-CGTase-H167C were optimized with response surface methodology. Maximum activity of β-CGTase-H167C was obtained with the medium containing 1.1% corn starch, 4.4% corn steep liquor, 1.1% peptone, 0.02% MgSO·7HO and 0.1% KHPO·3HO that were cultured with the initial pH 8.4, incubation temperature at 37.4 °C, with 5% inoculation size and shaking speed at 202 r/min. Under the optimal conditions, the activity of β-CGTase-H167C was up to 4355 U/mL, which is 1.93-fold in comparison with the initial activity. Our results established the promising culture strategy for the production of cyclodextrins by β-CGTase-H167C.

High-level production of γ-cyclodextrin glycosyltransferase in recombinant Escherichia coli BL21 (DE3): culture medium optimization, enzymatic properties characterization, and product specificity analysis

“…Hence, the hydrolysis activity of γ-CGTase plays an essential role in the manufacturing of γ-CD, even though the enzyme has been reported to primarily catalyze the transglycosylation reactions (cyclization, coupling, and disproportionation). The improved hydrolysis activity of CGTases depends on the significant expression vectors or the host cells, as well as the optimum culture media (Wang et al 2018b;Mahmud et al 2019;Upadhyay et al 2019). This is the first study to optimize the essential culture medium compositions of γ-CGTase to enhance the soluble expression and activity in E. coli BL21 (DE3) strain and explore its enzymatic properties to enhance the productivity and purity of γ-CD.…”

“…Under the optimum conditions, the maximum predicted γ-CGTase activity was found to be 53992.10 U mL −1 , which was 2.83-fold greater than the observed value in the culture medium before optimization. To the best of our knowledge, most of the known γ-CGTases produced by the wild microbial strains have exhibited extremely low hydrolysis activity or cyclization activity (Takada et al 2003;Hirano et al 2006;Goo et al 2014;Ji et al 2011;Wang et al 2018aWang et al , 2018b). In the current study, the experimental γ-CGTase hydrolysis activity (52680.86 ± 329.72 U mL −1 ) was significantly higher than the wild enzyme and the previously reported studies.…”

Enhanced production of questin by marine-derived Aspergillus flavipes HN4-13

“…Many useful metabolites can be produced by microbial fermentation (Cheng et al 2013;Dey et al 2018;Wang et al 2018;Zhao et al 2016b). Fermentation conditions depend on many variables, and the interactions among them are complicated.…”

Design of a Cyclodextrin Bioproduction Process Using Bacillus pseudofirmus and Paenibacillus macerans

“…The protocol described by Moriwaki et al [30] was adapted for this study. Briefly, a stock solution of PHE (3 mmol/L), in ethanol 95%, an aqueous buffer solution of sodium carbonate (0.6 mol/L) and sodium bicarbonate, and standard solutions of β-CD (2 mmol/L), in 20% (v/v) tris-HCl 50 mmol/L, 10% (v/v) CaCl 2 , 50 mmol/L, and water [25].…”

“…The application and study of alkalophilic Bacillus species for industrial production of extracellular enzymes is extensive due to their widespread presence, non-pathogenic nature, and high production capacity of extracellular enzymes. These enzymes, when easily isolated, exhibit specific characteristics such as activity and stability in extremely alkaline environments, and they often demonstrate activity or thermostability over a broad pH range compared to enzymes from neutralophiles [11,[23][24][25].…”