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oai:recercat.cat:2445/1770362025-12-05T09:34:50Zcom_2072_1057col_2072_478799col_2072_478917

Biological properties of poly-L-lysine/DNA complexes generated by cooperative binding of the polycation Liu, Ge Molas, Maria Grossmann, Gregory A. Pasumarthy, Murali Perales Losa, Carlos Cooper, Mark J. Hanson, Richard W. ADN Metabolisme Lisina DNA Metabolism Lysine We have evaluated the effect of NaCl concentration on the mode of binding of poly-L-lysine to DNA and the resulting structural and functional features of the condensed DNA particles using DNA precipitation, DNase I resistance, electron microscopy, and receptor-mediated gene transfer assays. At a high concentration of NaCl and in the presence of excess DNA, poly-L-lysine interacted with DNA cooperatively, fully condensing some of the DNA and leaving the rest of the DNA unbound. At low NaCl concentrations, poly-L-lysine molecules interacted with DNA in a noncooperative fashion, i.e. they bind randomly to the whole population of DNA molecules. Cooperative binding of poly-L-lysine to DNA occurred over a narrow range of NaCl concentrations, and the specific salt concentration depended on the length of the poly-L-lysine. The ability of condensed DNA to withstand digestion by DNase I was correlated with the structural features of the condensed DNA as determined by electron microscopy. Using our condensation procedure, cooperative binding of poly-L-lysine to DNA is a necessary prerequisite for the preparation of condensed DNA having a spherical shape and a diameter of 15-30 nm. Condensed DNA, containing galactosylated poly-L-lysine, was evaluated further for the extent and specificity of receptor-mediated gene transfer into HuH-7 human hepatoma cells via the asialoglycoprotein receptor. Efficient receptor-mediated transfection occurred only when condensed DNA complexes had a spherical shape with a diameter of 15-30 nm; asialofetuin, a natural ligand for the asialoglycoprotein receptor, inhibited this process by up to 90%. Our results support the importance of appropriate DNA condensation for the uptake and ultimate expression of DNA in hepatic cells. 2021-05-05T15:16:55Z 2021-05-05T15:16:55Z 2001-09-14 2021-05-05T15:16:56Z info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion 0021-9258 https://hdl.handle.net/2445/177036 174985 11438546 eng Reproducció del document publicat a: https://doi.org/10.1074/jbc.M105250200 Journal of Biological Chemistry, 2001, vol. 276, num. 37, p. 34379-34387 https://doi.org/10.1074/jbc.M105250200 (c) American Society for Biochemistry and Molecular Biology, 2001 info:eu-repo/semantics/openAccess 9 p. application/pdf American Society for Biochemistry and Molecular Biology Articles publicats en revistes (Ciències Fisiològiques)