Biofilms are a self‐organized, self‐structured community of microorganisms that are enclosed in a self‐produced polymeric matrix. Due to the intrinsic complexity of biofilms, their artificial preparation has remained a challenge despite practical usefulness as reservoirs for biocatalysts. Herein, autonomously generated artificial biofilms termed BIOMOSAIC (BioInspired, Oxygen‐induced Microbial Organization through Self‐Assembly at the air/liquid Interface as a Catalyst) films are reported. Similar to bacterial homing, a large number of cells are spontaneously recruited and immobilized into the film, which is autonomously generated at air/liquid interfaces by simple mixing of phenolic polyamine and tyrosinase‐expressing Pseudomonas fluorescens. BIOMOSAIC films are self‐regenerated, portable, reusable, and freestanding. They are multifunctional depending on types of gene expressions from the cells, catalyzing various reactions such as l‐DOPA production, dephosphorylation, and lipid hydrolysis. Furthermore, BIOMOSAIC film exhibits effective crude oil degradation properties, indicating that it can be a new self‐producing and self‐sustaining platform for bioremediation.