Kinetic inhibition of human salivary alpha-amylase by a novel cellobiose-containing tetrasaccharide.

OBJECTIVE

The aim of the study was to evaluate the inhibitory kinetics of a novel cellobiose-containing tetrasaccharide on human salivary alpha-amylase (HSA).

MATERIAL AND METHOD

Synthesis of cellobiose-containing tetrasaccharide was catalyzed by Paenibacillus sp. All CGTase using beta-CD as a donor and cellobiose as an acceptor under the optimal conditions. The reaction mixture was analyzed by HPLC and a cellobiose-containing tetrasaccharide obtained was studied for its inhibitory kinetics.

RESULTS

In vitro activity of human salivary alpha-amylase showed the optimum pH and temperature at 7.0 and 37 degrees C, respectively. The effects of metal ions, protective chemicals and saccharides on alpha-amylase activity, they were found that 10 mM concentration of CaCl2 and NaCl enhanced the enzyme activity. In contrast, the enzyme activity was significantly inhibited by 10 mM of HgCI2, alpha-cyclodextrin (alpha-CD) and synthetic cellobiose-containing tetrasaccharide. Chemicals often used as protective substance for enzyme such as beta-mercaptoethanol, EDTA or used as fungicide during enzyme purification (NaN3) had no effect on the activity of this enzyme. As a cellobiose-containing tetrasaccharide was shown to have a pronounce inhibition on alpha-amylase activity. Its inhibition kinetic was performed and found that cellobiose-containing tetrasaccharide was a competitive inhibitor with a Ki value of 7.89 microM.

CONCLUSION

Inhibition kinetic of a cellobiose-containing tetrasaccharide on alpha-amylase activity was competitive type with Ki value of 7.89 microM. In addition, these results will be a basic knowledge in controlling alpha-amylase actions that have influence on blood glucose level of trial animal and human further