PURIFICATION OF THE RUMINOCOCCUS-ALBUS ENDOGLUCANASE-IV USING A CELLULOSE-BINDING DOMAIN AS AN AFFINITY TAG

The gene encoding the single cellulose-binding domain II (CBD II) of C lostridium stercorarium xylanase A was fused to the egIV gene encoding endoglucanase IV (EGIV) from Ruminococcus albus. The fusion protein ( EGIV + CBDII) expressed in Escherichia coli can be readily purified fr om the cell-free extract of E. coli in a single step using the affinit y of CBD to cellulose. The purified enzyme was cleaved into two moieti es, i.e. the catalytic-domain and CBD, at a specific site in the linke r region by partial digestion with trypsin at 4 degrees C. This result indicates that this CBD belonging to family VI of CBD families can be used as an affinity tag for purification of the recombinant protein.