Kinetics of the urea–urease clock reaction with urease immobilized in hydrogel beads

Feedback driven by enzyme catalyzed reactions occurs widely in biology and has been well characterized in single celled organisms such as yeast. There are still few examples of robust enzyme oscillators in vitro that might be used to study nonlinear dynamical behavior. One of the simplest is the urea–urease reaction that displays autocatalysis driven by the increase in pH accompanying the production of ammonia. A clock reaction was obtained from low to high pH in batch reactor and bistability and oscillations were reported in a continuous flow rector. However, the oscillations were found to be irreproducible and one contributing factor may be the lack of stability of the enzyme in solution at room temperature. Here, we investigated the effect of immobilizing urease in thiol-poly(ethylene glycol) acrylate (PEGDA) hydrogel beads, prepared using emulsion polymerization, on the urea–urease reaction. The resultant mm-sized beads were found to reproduce the pH clock and, under the conditions employed here, the stability of the enzyme was increased from hours to days.