Hybrid spheres containing peptides and gold nanoparticles have been simultaneously synthesized in water using AG4 (NPSSLFRYLPSD) peptides that acted as a reducing agent to guide the nucleation and growth of gold nanoparticles and a precursor to form sphere-like structure by self-assembly where the size of hybrid spheres is precisely controlled by adjusting the operating temperature. The self-assembled peptide spheres remain stable even after selective removal of the gold nanoparticles by iodide etching. The amino acids containing the aromatic functional group in the peptide sequence significantly affect the construction of sphere structures. The surface of gold nanoparticles containing hybrid spheres has been functionalized using the thiol group linked to biomolecules. The ability to synthesize nanoparticle and self-assembled peptide structures with controlled size and composition in an environmental benign way will allow us to fabricate a new class of multifunctional organic-inorganic hybrid superstructures for various biomedical and electronic applications.