Purification and the effect of manganese ions on the activity of carboxymethylcellulases from Aspergillus niger and Cellulomonas biazotea

Carboxymethylcellulases (CMCases) from Aspergillus niger and Cellulomonas biazotea were purified by a combination of ammonium sulfate precipitation, anion-exchange and gel-filtration chromatography with a 12- and 9-fold increase in the purification factor. The native and subunit molar mass of CMCase from A. niger were 40 and 25-57 kDa, respectively, while those from C. biazotea were 23 and 20-30 kDa, respectively. Low concentrations of Mn2+ activated the enzymes from both organisms (mixed activation) with apparent activation constants of 0.80 and 0.45 mmol/L of CMCases from A. niger and C. biazotea, respectively, while at higher CMC concentrations Mn2+ inhibited the enzymes (mixed and partial uncompetitive inhibition). The reason for this complex behavior is that more than one Mn2+ bind to the same enzyme form with the apparent average inhibition constants of 2.7 and 1.3 mmol/L for CMCases from A. niger and C. biazotea, respectively.