Perales J C, Grossmann G A, Molas M, Liu G, Ferkol T, Harpst J, Oda H, Hanson R W
Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4935, USA.
J Biol Chem. 1997 Mar 14;272(11):7398-407. doi: 10.1074/jbc.272.11.7398.
Electrostatic binding of polycations or basic polypeptides to the DNA phosphate backbone has been previously described as a one-step process which results in uncontrolled aggregation and precipitation of the DNA in solution. We describe here a multistep process in which the condensation of DNA in the presence of poly-L-lysine can be controlled to produce particles of discrete size and shape suitable for receptor-mediated gene transfer in vivo and in vitro. The first step in this process involves the gradual accretion of poly-L-lysine onto the DNA phosphate backbone, until charges are neutralized. The addition of poly-L-lysine to a concentrated solution of DNA in this fashion prevents intermolecular aggregation of the DNA, presumably by promoting the formation of a nucleus of condensation along the length of each DNA molecule. The second stage of the process involves adjusting the ionic strength of the solvent to facilitate the solubilization of compact DNA.poly-L-lysine complexes. Several physical and biochemical parameters have been studied and correlated with the efficacy of DNA/ligand-poly-L-lysine particles in transferring genes to the liver of adult animals by receptor-mediated endocytosis.
多阳离子或碱性多肽与DNA磷酸骨架的静电结合此前被描述为一个一步过程，该过程会导致溶液中的DNA无控制地聚集和沉淀。我们在此描述一个多步过程，其中在聚-L-赖氨酸存在下DNA的凝聚可得到控制，从而产生大小和形状离散的颗粒，适合在体内和体外进行受体介导的基因转移。该过程的第一步涉及聚-L-赖氨酸在DNA磷酸骨架上逐渐积累，直至电荷被中和。以这种方式向浓DNA溶液中添加聚-L-赖氨酸可防止DNA分子间聚集，推测这是通过沿每个DNA分子的长度促进凝聚核的形成来实现的。该过程的第二阶段涉及调节溶剂的离子强度，以促进紧密的DNA-聚-L-赖氨酸复合物的溶解。已经研究了几个物理和生化参数，并将其与DNA/配体-聚-L-赖氨酸颗粒通过受体介导的内吞作用将基因转移到成年动物肝脏的效率相关联。
