Improved gene expression using low molecular weight peptides produced from protamine sulfate

DNA condensation plays a key role in non-viral gene delivery by affecting gene transfection, nuclear targeting, and eventual gene expression efficiency. Theoretically, a DNA condenser with the appropriate DNA condensation ability but without affecting DNA dissociation from DNA condensates inside the cytoplasm should be a perfect carrier for gene delivery. Protamine is a natural DNA condensation agent and has been widely used in gene delivery. In this work, protamine was selectively digested enzymatically to produce low molecular weight protamine fragments (LMWPs) of various lengths and amino acid compositions. The DNA condensation ability and gene transfection efficiency of these LMWP peptides were tested. Compared to protamine, all the LMWP peptides showed lower DNA binding strength. However, some LMWP peptides demonstrated excellent DNA condensation ability and could form very compact DNA condensates with small particle size (approximately 100 nm). More interestingly, LMWP peptide-mediated in vitro gene delivery showed prolonged (up to 12 days) gene expression. Results from this study suggest that designing DNA condensers with appropriate and tunable DNA binding strengths and condensation abilities would be an effective means to improve gene expression and thus gene therapy efficiency. Since LMWP peptides have low immunogenicity, they would be safer than protamine for use in gene therapies.