Affinity partitioning of a Cellulomonas fimi β-mannanase with a mannan-binding module in galactomannan/starch aqueous two-phase system

A new approach in affinity separations was studied by partitioning of Cellulomonas fimi β-mannanase (EC 3.2.1.78) containing a mannan-binding module in galactomannan/hydroxypropyl starch aqueous two-phase system. Comparison was made with a truncated version of C. fimi β-mannanase which lacked the mannan-binding module. Results showed that affinity partitioning of the β-mannanase was achieved due to biospecificity of the mannan-binding module towards the top phase containing galactomannan. Experiments were conducted at pH 8 to prevent enzyme degradation of the phase containing galactomannan. Removal of the top phase polymer was accomplished by β-mannanase degradation allowed by shifting to the optimal pH 6. In the combination with the genetic fusion of any given protein to the mannan-binding module, the results envision a general procedure for primary affinity recovery of such fusion proteins.