Chemical glycosidation of trypsin with O-carboxymethyl-poly-beta-cyclodextrin: catalytic and stability properties.
The polysaccharide O-carboxymethyl-poly-beta-cyclodextrin was synthesized (molecular mass 13,000 Da, 40% carboxy groups) and attached to the surface of bovine pancreatic trypsin. The resulting neoglycoenzyme retained high proteolytic and esterolytic activity and contained approx. 1.0 mol of polymer/mol of enzyme. The optimum temperature for trypsin activity was increased by 10 degrees C after this transformation. Thermostability of the polymer-enzyme complex was increased by about 14 degrees C over 10 min incubation. The conjugate was also more resistant to thermal inactivation at different temperatures, ranging from 45 to 60 degrees C, demonstrating the influence of supramolecular and polymer-protein electrostatic interactions on trypsin thermostabilization. Additionally, the conjugate was 36-fold more resistant to the action of the anionic surfactant SDS. This modification also protected the enzyme from autolysis at alkaline pH.
Villalonga
Maria L
ML
Enzyme Technology Group, Center for Biotechnological Studies, University of Matanzas, Autopista a Varadero km 3 1/2, Matanzas, C.P. 44740, Cuba.
Reyes
Gretel
G
Fragoso
Alex
A
Cao
Roberto
R
Fernández
Leyden
L
Villalonga
Reynaldo
R
D000818:Animals; D002266:Carboxymethylcellulose Sodium / Q000737:chemistry*; D002384:Catalysis; D002417:Cattle; D004789:Enzyme Activation; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D010179:Pancreas / Q000201:enzymology; D011485:Protein Binding; D011489:Protein Denaturation; D055672:Static Electricity; D013696:Temperature; D014357:Trypsin / Q000138:chemical synthesis / Q000737:chemistry*; D047392:beta-Cyclodextrins / Q000737:chemistry*
D046911:Macromolecular Substances; D047392:beta-Cyclodextrins; D014357:Trypsin; D002266:Carboxymethylcellulose Sodium