Two-dimensional (2D) nanomaterials have received increasing interest for many applications such as biomedicine and nanotechnology. Here, we report a facile strategy to prepare highly flexible 2D crystalline nanosheets with only ∼6 nm thickness from poly­(ethylene glycol)-block-poly­(N-octylglycine) (PEG-b-PNOG) diblock copolymer in high yield. To our best knowledge, this is the first report of free-floating, 2D extended nanosheets from polypeptoid-based block copolymers. The faceted nanostructures are achieved from hierarchical self-assembly through a sphere-to-cylinder-to-nanosheet transition pathway. The preliminary assembled spheres can behave like a fundamental packing motif to spontaneously stack into a 2D lattice via an intermediate cylinder structure, driven by crystallization of PNOG domains. The nanosheet formation process follows theoretical model for morphology development of crystalline block copolymers in selective solvents. Particularly remarkable is that we obtained the hierarchical nanostructure from synthetic block copolymers through a multiple-step strategy mimetic to protein crystallization. This is fairly distinct from the previously reported crystalline nanosheets. The ability to efficiently create 2D crystals from synthetic polymers by spontaneous assembly will enable new generations of bioinspired nanomaterials for a variety of potential applications in biomedicine and nanotechnology.