Two-step sequential reaction catalyzed by layer-by-layer assembled urease and arginase multilayers

Layer-by-layer (LbL) self-assembly was used to build multi-component thin films containing polyions and enzymes: urease (Ur), and arginase (Ar). These multilayers were shown to participate catalytically in a two step sequential decomposition Of L-arginine to urea and subsequently to ammonia. Deposition of multilayers was characterized with quartz crystal microbalance (QCM). The following enzyme monolayer stacking architectures were studied: (Ur/ PDDA)(n), (Ar/PDDA)(n), (Ar/PDDA/Ur/PDDA)(n), and (Ur/PDDA/Ar/PDDA)(n), where n = 1-4, PDDA and PSS are, respectively, poly(dimethyldiallyl ammonium chloride) and sodium poly(styrene sulphonate). The layer growth on gold electrode resonator was monitored with QCM and the catalytic activity of the enzyme multilayers was studied with UV-Vis spectroscopy, using a pH-sensitive dye (bromocresol purple). Dependence of film catalytic activity was studied as a function of position and number of the enzyme layers. The assembly with layer sequence {PDDA/PSS/PDDA+ (Ur/PDDA/Ar/PDDA)(2)} showed the highest catalytic activity. The order of enzyme multilayers in this architecture corresponds to the two-step sequential reaction: L-arginine --> urea --> ammonia. The assembly was then fabricated on the tip of a commercial ammonia electrode and detection Of L-arginine was demonstrated at physiologically significant concentrations. (C) 2003 Elsevier Science B.V. All rights reserved.