UREASE FROM THE SOIL BACTERIUM BACILLUS-PASTEURII - IMMOBILIZATION ONCA-POLYGALACTURONATE

Urease purified from the soil bacterium Bacillus pasteurii was adsorbe d and immobilized on a preformed network of Ca-polygalacturonate, a su bstrate which has a similar composition and morphology to the mucigel present at the root-soil interface. The adsorption proceeded with an e ssentially quantitative yield, and the immobilized enzyme showed no de crease of specific activity with respect to the free enzyme. The depen dence of urease adsorption on NaCl concentration suggested that the en zyme is bound to the carrier gel through electrostatic interactions. T he immobilized enzyme showed increased stability, with respect to the free enzyme, with increasing time or temperature, and in the presence of proteolytic enzymes. The pH activity profile revealed that the adso rbed enzyme showed no change in the optimum pH (8.0), but it was more active than the free form in the pH range 5-8. The Michaelis-Menten ki netic parameters V-max and K-m were measured for the free (V-max = 196 0 +/- 250 units ml(-1); K-m = 235 +/- 20 mM) and immobilized (V-max = 1740 +/- 185 units ml(-1); K-m = 315 +/- 25 mM) urease. The substantia l similarity of V-max in the two cases suggests that there were no con formational changes involving the active site upon enzyme immobilizati on, while substrate partitioning effects between the bulk solution and the micro-environment surrounding the immobilized enzyme must be oper ating so as to partly increase its K-m. These results suggest that bac terial urease present in plant root mucigel plays a large role in the mobilization of urea N. Its activity is in fact significantly mantaine d and protected by immobilization on hydrophilic gels such as those pr oduced by root exudates. Copyright (C) 1996 Published by Elsevier Scie nce Ltd