With the synthesis and functionalization of membranes for selective separations, reactivity, and stimuli responsive behavior arises new and advanced opportunities. The integration of bio-based channels is one of these advancements in membrane technologies. By a layer-by-layer (LbL) assembly of polyelectrolytes, outer membrane protein F trimers (OmpF) or “porins” from Escherichia coli with a central pore of ~2 nm diameter at its opening and ~0.7 × 1.1 nm at its constricted region are immobilized within the pores of poly(vinylidene fluoride) microfiltration membranes, as opposed to traditional ruptured lipid bilayer or vesicles processes. These OmpF-membranes demonstrate selective rejections of non-charged organics over ionic solutes, allowing the passage of salts up to 2 times higher than traditional nanofiltration membranes starting with rejections of 84% for 0.4–1.0 kDa organics. The presence of charged groups in OmpF membranes also leads to pH-dependent salt rejection through Donnan exclusion. These OmpF-membranes also show exceptional durability and stability, delivering consistent and constant permeability and recovery for over 160 h of operation. Characterization of solutions containing OmpF, and membranes were conducted during each stage of the process, including detection by fluorescence labelling (FITC), zeta potential, pH responsiveness, flux changes, and rejections of organic-inorganic solutions.