Kinetics of β-cyclodextrin production by cyclodextrin glucanotransferase from Bacillus megaterium

Abstract

The kinetics of β-cyclodextrin production by cyclodextrin glucanotransferase from Bacillus megaterium was investigated using two different approaches. By studying the initial rate of the enzyme reaction a strong inhibition of CGTase by the main product of the cyclization reaction β-CD was revealed. The type of inhibition was determined to be mixed competitive and uncompetitive. The value of the uncompetitive inhibition constant (Kiu=33.30 mg/mL) was much higher than the value of the competitive inhibition constant (Kic=4.21 mg/mL), which indicated a predominance of the competitive part of the mixed type inhibition. The dynamics of β-CD formation and the corresponding starch degradation was investigated at real process conditions with different initial substrate concentrations. The enzyme reaction rate was maximal at the beginning of the process when the substrate current concentration was maximal. Depletion of the substrate led to a corresponding decrease in rate values. Another reason for the decrease in the enzyme rate was the accumulation of the product β-CD, which inhibited CGTase. The results for the dynamics of beta CD production were used for investigation of the kinetics of the process at real process conditions. In a real CD production process, CGTase showed a change in the type of inhibition by β-CD. The type of inhibition was determined to be competitive, with inhibition constant Ki=6.13 mg/mL. A probable reason for the change of the type of inhibition at real process conditions was the change in the substrate and product concentrations during the process.