CONSTRUCTION AND CHARACTERIZATION OF A CHIMERIC BETA-GLUCOSIDASE

The amino acid sequences of beta-glucosidases from Cellvibrio gilvus a nd Agrobacterium tumefaciens show significant similarity in most of th e parts. However, the pH/temperature optima and stabilities of the two enzymes are quite different. C. gilvus beta-glucosidase exhibits an o ptimum pH of 6.2-6.4 and temperature of 35 degrees C, whereas the corr esponding values for A. tumefaciens are 7.2-7.4 and 60 degrees C respe ctively. To analyse these properties further, a chimeric beta-glucosid ase was constructed by replacing a segment from the C-terminal region of C. gilvus beta-glucosidase gene with that of A. tumefaciens. The pa rtially purified chimeric enzyme was characterized with respect to pH/ temperature activity and stability and substrate affinity. Our results suggest that C-terminal segment(s) might be important in beta-glucosi dase specificity, and shuffling of even a small segment of gene in thi s region might significantly alter or improve the enzymic properties s uch as thermal stability.