Characterization of amylose affinity membranes through adsorption of pure recombinant MBP-intein-CBD: equilibrium and kinetics data

摘要

Pure MBP-intein-CBD has been used as model protein in adsorption stages on amylose affinity membranes. Equilibrium data have been obtained and fitted to the Langmuir isotherm, and the model parameters have been estimated. The influence of flowrate on adsorbed protein has been investigated using a continuous flow system. Steady state data have been then compared with the equilibrium curve previously obtained. The kinetic constants of the binding and desorption reactions have been estimated using a kinetic model accounting for both adsorption and desorption reactions. Mass transfer effects within the liquid phase have been found negligible. Values of kinetic parameters have been obtained in the operative conditions of both adsorption and elution steps. The influence of NaCl concentration on the values of k_1 and k_2 can be roughly estimated. The knowledge of these parameters, together with the maximum binding capacity allows for the scale-up design of an affinity membrane chromatographic column.