A simple and effective method is described for the fabrication of robust zeolitic membranes with three-dimensional (3D) interconnected macroporous structures. The membranes were prepared by electrostatically seeding mesoporous silica sphere (MSS) self-assembled films with silicalite-1 nanoparticles, followed by hydrothermal treatment. The membrane thickness, which is determined by the MSS film thickness, can be easily adjusted from tens to hundreds of micrometers by varying the concentration of the MSS dispersion and the solution volume: Biomacromolecule-functionalized macroporous zeolitic membrane bioreactors were subsequently prepared-via the layer-by-layer. (LbL) electrostatic assembly of polyelectrolytes and enzyme (catalase) on the 3D macroporous membranes. The enzyme-modified membranes with interconnected macroporous structures display enzyme loading amounts and activities that are one order of magnitude higher than corresponding 3D zeolite films with closed macropores, and approximately three orders of magnitude higher than their non-porous planar film counterparts assembled on silica substrates. The enzyme loadings and activities were found to be approximately linearly dependent on the thicknesses of the membranes. Furthermore, the immobilized enzyme exhibits enhanced reaction stability in comparison with enzyme in bulk solution. These membranes are potentially useful for separations as they could be used to simultaneously perform reaction and separation, steps.