DNA and Peptide Functionalized Gold Nanoparticles for Biological Imaging and Transfection

This dissertation focuses on the development of DNA and peptide functionalized gold nanoparticles. Three nanoparticle systems have been developed:: 1) nucleic acid-directed self-assembly of a gold nanoparticle PET imaging agent;: 2) cationic peptide functionalized gold nanoparticles;: 3) gold nanoparticle templated peptide nanoshells. The first functionalized gold nanoparticles were constructed by assembling ODN: oligodeoxynucleotide)-derivatized gold nanoparticles with functionalized complementary PNAs: peptide nucleic acids). The PNAs were conjugated with DOTA for chelating chelating 64Cu for PET imaging, PEG for conferring stealth properties, and Cy5 for fluorescent imaging. The resulting functionalized nanoparticles showed good stability both in vitro and in vivo and had biodistribution properties expected for a PEGylated gold nanoparticle rather than that for the functionalized PNAs used in the construction. The second nanoparticle system was arginine-rich peptide and lysine-rich peptide functionalized cationic gold nanoparticles as transfection agents. A series of cysteine containing oligoarginine was synthesized by automated solid phase Fmoc synthesis. chloroauric acid was reduced in the presence of the peptides. The resulting gold nanoparticles were around 13 nm in diameter with zeta potential around 29 mV. Compare to other reported cationic gold nanoparticles, the arginine-rich gold nanoparticles were highly stable in up to 0.6 M NaCl solutions and up to pH 8.5 in non-phosphate containing buffers. In phosphate buffers, however, the nanoparticles started to aggregate at above pH 6, though aggregation was reversible by lowering the pH. The strong affinity between arginine and phosphate groups also resulted in the nanoparticle\u27s strong ODN binding ability. Despite of the reported cytotoxicity of oligoarginines, the arginine-rich gold nanoparticles showed no cytotoxicity even at a very high concentration. An improved assembly approach was also investigated. Cationic peptides were exchanged with cetyltrimethylammonium bromide: CTAB) on gold nanoparticles fabricated by seeded growth method. This approach allowed quick assembly of a library of different peptide on the gold surface. The approach was validated by synthesizing three cationic gold nanoparticles, R13C-AuNP, K10Y2C-AuNP and R13C/K10Y2C-AuNP. All of the nanoparticles retained the monodispersity of the original CTAB·AuNP colloid. Their stability and ODN binding ability showed distinct difference due to their different surface composition. All of the colloids were proved to be non-toxic at high concentration but further optimization is needed in the transfection efficiency. The third nanoparticle system was developed based on the ligand exchange approach. A cysteine containing lysine-rich peptide was deposited on CTAB·AuNP. Then the lysines were crosslinked by a biodegradable crosslinker, Sulfo-EGS: Ethylene glycol bis[sulfosuccinimidylsuccinate]), at various peptide/crosslinker ratio. The gold core was then removed by potassium cyanide. The resulting peptide nanoshells were about 40 nm in diameter, compare to the 15 nm diameter before the gold core was removed. Crosslinking also resulted in dramatic decrease of the ODN binding ability of the nanoparticle and the nanoshells. This approach showed great potential of making a combinatorial library of biodegradable nanostructures quickly for a broad range of applications