Investigation of DNA spectral conformational changes and polymer buffering capacity in relation to transfection efficiency of DNA/polymer complexes.
OBJECTIVE
The relation between transfection efficiency of DNA/polymer complexes and DNA conformational alterations is investigated. The buffering capacity of several synthetic polymers is also studied to relate their performance in transfection efficiency.


METHODS
The cationic polymer/DNA interaction was evaluated by measuring the alteration of DNA secondary structures in solution before and after the addition of polymer with ATR-FTIR technique. The degree of protonation in aqueous cationic polymers is varied upon pH and different structures. A polymer capable of protonation acts like a proton sponge to react with H+ in titration with HCl. This characteristic was evaluated in relation to transfection efficiency because the capacity would help the release of endocytotic DNA from endosome/lysosome on its way to expression.


RESULTS
IR results show that the antisymmetric PO2- vibration of DNA (at 1224 cm-1) shifts toward lower frequencies in complexation with PEI or PLLys (these polymers are able to transfect DNA). By contrast, the antisymmetric PO2- vibration of DNA in presence of PDAMA or dextran (these polymers are poor in DNA transfection) shows a shifting to higher frequencies or no alteration was observed. Interestingly, the polymers with best performance in transfection efficiency are in this order: PEI>PDMAEMA>PLLys>PDAMA>dextran which is in the same order as their polymer buffering capacity. These facts indicate polymers possessing better buffering capacity could result in higher transfection efficiency. Also, we have demonstrated in this paper that the alteration in the antisymmetric PO(2)(-) stretching vibration in the IR spectra is sensitive to the binding of cationic polymers with DNA and the transfection efficiency of the formed complexes. These findings are useful for the developing polymer-based gene delivery systems with better in vitro and in vivo performance. NOVELTY OF THE WORK: this work establishes a rational relation between transfection efficiency of DNA/polymer complexes and DNA conformational alterations. Also, the buffering capacity of polymers is found to be closely related to their performance in transfection efficiency.
Cherng
Jong Yuh
JY
Department of Chemistry and Biochemistry, National Chung Cheng University, 168, University Rd. Chia-Yi 621, Taiwan, ROC. jycherng@yahoo.com
D000818:Animals; D002021:Buffers; D019556:COS Cells; D002412:Cations / Q000737:chemistry; D002522:Chlorocebus aethiops; D006224:Cricetinae; D004247:DNA / Q000737:chemistry*; D003911:Dextrans / Q000737:chemistry; D018014:Gene Transfer Techniques; D006863:Hydrogen-Ion Concentration; D008689:Methacrylates / Q000138:chemical synthesis / Q000737:chemistry; D009757:Nylons / Q000138:chemical synthesis / Q000737:chemistry; D010957:Plasmids; D011094:Polyethyleneimine / Q000737:chemistry; D011107:Polylysine / Q000737:chemistry; D011108:Polymers / Q000138:chemical synthesis / Q000737:chemistry*; D011109:Polymethacrylic Acids / Q000138:chemical synthesis / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared; D014162:Transfection / Q000379:methods
D002021:Buffers; D002412:Cations; D003911:Dextrans; D008689:Methacrylates; D009757:Nylons; D011108:Polymers; D011109:Polymethacrylic Acids; C407037:poly(2-(dimethylamino)ethyl methacrylate); C521988:poly(2-methylacrylic acid 2-((2-(dimethylamino)ethyl)methylamino)ethyl ester); D011107:Polylysine; D011094:Polyethyleneimine; D004247:DNA; C427203:calf thymus DNA