title: Low molecular weight protamine as an efficient and nontoxic gene carrier: in vitro study.
abstract: BACKGROUND
  The structural similarity between low molecular weight protamine (LMWP), prepared by enzymatic digestion of protamine, and HIV-TAT protein transduction peptide suggested the feasibility of LMWP as an efficient carrier for delivering therapeutic genes while alleviating the cytotoxicity of currently employed gene carriers.
  
  
  METHODS
  LMWP was prepared by enzymatic digestion of protamine with thermolysine. The prepared LMWP peptide and TAT peptide, as well as their complexes with plasmid DNA (pDNA), were examined for cellular uptake behaviors by using confocal microscopy and flow cytometry. The complexation of pDNA and LMWP was monitored by gel retardation test as well as size and zeta potential measurements, and was then further assessed by DNase I protection assay. The transfection efficiency of pDNA/LMWP was examined by varying the pDNA content and charge ratio in the complex, and then compared with that of pDNA/PEI. Cytotoxicity induced by pDNA/LMWP and pDNA/PEI was also examined.
  
  
  RESULTS
  Prepared LMWP showed similar transcellular localization behavior and kinetics to those of TAT, and efficiently transferred the pDNA into nucleus and cytoplasm in a short time period. The size and zeta potential of the pDNA/LMWP complex were 120 nm and 30 mV, respectively, which were adequately suitable for cellular uptake. After forming the complex, LMWP appeared to effectively protect pDNA against DNase I attack. The pDNA/LMWP complex showed significantly enhanced gene transfer than both naked pDNA and the pDNA/PEI complex, while exhibiting a markedly reduced cytotoxicity than that of the pDNA/PEI complex.
  
  
  CONCLUSIONS
  The present study suggested that LMWP could be a useful and safe tool for enhancing delivery of bioactive molecules and therapeutic DNA products into cells when applied in gene therapy.
authors.affiliation: College of Pharmacy, The University of Michigan, Ann Arbor, MI 48109, USA.
authors.forename: Yoon Jeong
authors.initials: YJ
authors.lastname: Park
authors.forename: Jun Feng
authors.initials: JF
authors.lastname: Liang
authors.forename: Kyung Soo
authors.initials: KS
authors.lastname: Ko
authors.forename: Sung Wan
authors.initials: SW
authors.lastname: Kim
authors.forename: Victor C
authors.initials: VC
authors.lastname: Yang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002470:Cell Survival; D004247:DNA / Q000378:metabolism; D015696:Gene Products, tat / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D006801:Humans; D046911:Macromolecular Substances; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011479:Protamines / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use*; D014162:Transfection
chemical_list: D015696:Gene Products, tat; D046911:Macromolecular Substances; D010455:Peptides; D011479:Protamines; D004247:DNA