Two-dimensional nanostructures offer significant promise as components for the construction of functional biomaterials. However, the controllable fabrication of these structures remains a challenge. Ideally, one desires to control the composition, structure, and surface functionality of the resultant materials with precision, in order to tailor properties for a particular application and minimize the unintended side effects. We recently reported the synthesis of triple-layer nanosheets from template-driven assembly of a negatively charged collagen-mimetic peptide CP on a preassembled nanosheet of a positively charged collagen-mimetic peptide CP [1]. This process enabled the fabrication of nanosheets of defined composition, internal structure, and surface chemistry using a modified layer-by-layer approach. Herein, we describe the synthesis and purification procedures for these two 45-mer peptides, CP and CP , and guidelines for the directed assembly of triple-layer structures, along with routine methods of structural analysis.
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