GENERATION OF A PH GRADIENT IN AN IMMOBILIZED ENZYME-SYSTEM

Several examples of two-step sequential reactions exist where, because of the poor equilibrium conversion by the first reaction, it is desir able to conduct the two reactions simultaneously. In such a scheme, th e product of the first reaction is continuously removed by the second reaction, thus not allowing the first reaction to approach chemical eq uilibrium. Therefore, the first reaction is allowed to proceed in the desired direction at an appreciable rate. However, in many biochemical applications where enzyme catalysts are involved, the enzyme's activi ties are strong functions of pH. Where the pH optima of the first and second reaction differ by three to four units, the above reaction sche me would be difficult to implement, In these cases, the two reactions can be separated by a thin permeable membrane across which the desired pH gradient is maintained. In this article, it was shown, both by the ory and experiment, that a thin, flat membrane of immobilized urease c an accomplish this goal when one face of the membrane is exposed to th e acidic bulk solution (pH(b) = 4.5) containing a small quantity of ur ea (0.01 M). In this particular case, the ammonia that was produced in the membrane consumed the incoming hydrogen ions and thus maintained the desired pH gradient. Experimental results indicate that with suffi cient urease loading, the face of the membrane opposite to the bulk so lution could be maintained at a pH that would allow many enzymes to re alize their maximum activities (almost-equal-to 7.5). It was also foun d that this pH gradient could be maintained even in the presence of a buffer, which greatly enhances the transport of protons into the membr ane. (C) 1993 John Wiley & Sons, Inc.