Abstract
The interaction of two large beta-galactosidases (from
Escherichia coli and from
Thermus sp.) with tailor-made anion exchangers was studied. Using lowly activated supports (e.g., containing 2–3
μmol of ionised groups per wet gram of support), large proteins selectively adsorbed and easily desorbed (e.g., using 200
mM of NaCl), giving highly purified proteins. However, these supports cannot be used to immobilize the enzymes for industrial use, because the weak adsorption.
On the other hand, these large proteins strongly adsorb on very highly activated supports (e.g., containing 40
μmol of ionic groups per wet gram of 4 BCL agarose). Thus, these supports may be not valid for large protein purification, but may be very suitable for immobilization of these proteins. Using high ionic strength (e.g., 300
mM NaCl), large proteins still may be adsorbed on these supports, while only around 20% of total proteins adsorb, permitting some purification of the large proteins but not a total one. Moreover, adsorption under these conditions increase the adsorption strength (now there are not desorption even using 800
mM NaCl).
Thus, the purification and the strong reversible immobilization of both beta-galactosidases were performed in a very simple two-step process. The large proteins can be directly adsorbed on these supports after desorption (at 200
mM of NaCl) from poorly activated supports. Furthermore, adsorption on very highly activated supports promotes a significant thermal stabilization of both enzymes, mainly in dissociations conditions.