Left-handed gold nanoparticle double helices were prepared using a new method that allows simultaneous synthesis and assembly of discrete nanoparticles. This method involves coupling the processes of peptide self-assembly of and peptide-based biomineralization of nanoparticles. In this study, AYSSGAPPMPPF (PEPAu), an oligopeptide with an affinity for gold surfaces, was modified with an aliphatic tail to generate C12-PEPAu. In the presence of buffers and gold salts, amphiphilic C12-PEPAu was used to both control the formation of monodisperse gold nanoparticles and simultaneously direct their assembly into left-handed gold nanoparticle double helices. The gold nanoparticle double helices are highly regular, spatially complex, and they exemplify the utility of this methodology for rationally controlling the topology of nanoparticle superstructures and the stereochemical organization of discrete nanoparticles within these structures.