ABSTRACT
Despite considerable interest and investigations on cationic lipid-DNA complexes, reports on lipid-RNA interaction are very limited. In contrast to lipid-DNA complexes where lipid binding induces partial B to A and B to C conformational changes, lipid-tRNA complexation preserves tRNA folded state. This study is the first attempt to investigate the binding of cationic lipid with transfer RNA and the effect of lipid complexation on tRNA aggregation and condensation. We examine the interaction of tRNA with cholesterol (Chol), 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), dioctadecyldimethylammoniumbromide (DDAB) and dioleoylphosphatidylethanolamine (DOPE), at physiological condition, using constant tRNA concentration and various lipid contents. FTIR, UV-visible, CD spectroscopic methods and atomic force microscopy (AFM) were used to analyze lipid binding site, the binding constant and the effects of lipid interaction on tRNA stability, conformation and condensation. Structural analysis showed lipid-tRNA interactions with G-C and A-U base pairs as well as the backbone phosphate group with overall binding constants of K(Chol) = 5.94 (+/- 0.8) x 10(4) M(-1), K(DDAB) = 8.33 (+/- 0.90) x 10(5) M(-1), K(DOTAP) = 1.05 (+/- 0.30) x 10(5) M(-1) and K(DOPE) = 2.75 (+/- 0.50) x 10(4) M(-1). The order of stability of lipid-tRNA complexation is DDAB > DOTAP > Chol > DOPE. Hydrophobic interactions between lipid aliphatic tails and tRNA were observed. RNA remains in A-family structure, while biopolymer aggregation and condensation occurred at high lipid concentrations.
Subject(s)
Lipids/chemistry , RNA, Transfer/chemistry , Cations/chemistry , Cholesterol/chemistry , Circular Dichroism , Fatty Acids, Monounsaturated/chemistry , Hydrophobic and Hydrophilic Interactions , Microscopy, Atomic Force , Nucleic Acid Conformation , Phosphates/chemistry , Phosphatidylethanolamines/chemistry , Quaternary Ammonium Compounds/chemistry , RNA, Transfer/ultrastructure , Spectroscopy, Fourier Transform InfraredABSTRACT
Complexes of cationic liposomes with DNA are promising tools to deliver genetic information into cells for gene therapy and vaccines. Electrostatic interaction is thought to be the major force in lipid-DNA interaction, while lipid-base binding and the stability of cationic lipid-DNA complexes have been the subject of more debate in recent years. The aim of this study was to examine the complexation of calf-thymus DNA with cholesterol (Chol), 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), dioctadecyldimethylammoniumbromide (DDAB) and dioleoylphosphatidylethanolamine (DOPE), at physiological condition, using constant DNA concentration and various lipid contents. Fourier transform infrared (FTIR), UV-visible, circular dichroism spectroscopic methods and atomic force microscopy were used to analyse lipid-binding site, the binding constant and the effects of lipid interaction on DNA stability and conformation. Structural analysis showed a strong lipid-DNA interaction via major and minor grooves and the backbone phosphate group with overall binding constants of K(Chol) = 1.4 (+/-0.5) x 10(4) M(-1), K(DDAB) = 2.4 (+/-0.80) x 10(4) M(-1), K(DOTAP) = 3.1 (+/-0.90) x 10(4) M(-1) and K(DOPE) = 1.45 (+/- 0.60) x 10(4) M(-1). The order of stability of lipid-DNA complexation is DOTAP>DDAB>DOPE>Chol. Hydrophobic interactions between lipid aliphatic tails and DNA were observed. Chol and DOPE induced a partial B to A-DNA conformational transition, while a partial B to C-DNA alteration occurred for DDAB and DOTAP at high lipid concentrations. DNA aggregation was observed at high lipid content.
Subject(s)
DNA/chemistry , Lipids/chemistry , Cations/chemistry , Cholesterol/chemistry , Circular Dichroism , DNA/ultrastructure , Fatty Acids, Monounsaturated/chemistry , Hydrophobic and Hydrophilic Interactions , Microscopy, Atomic Force , Nucleic Acid Conformation , Phosphates/chemistry , Phosphatidylethanolamines/chemistry , Quaternary Ammonium Compounds/chemistry , Spectroscopy, Fourier Transform InfraredABSTRACT
The anticancer platinum (Pt) drugs exert their antitumor activity by direct or indirect Pt-DNA binding. It has been shown that Pt drugs can induce major DNA damage and minor RNA damage during cancer treatment. A recent report showed that a new anticancer estradiol-Pt(II) hybrid molecule (CD-37) binds DNA bases indirectly, while being more effective than cis-diaminedichloroplatinum(II) (cisplatin) against several types of cancer. In this report, we examine the bindings of CD-37 and cisplatin drugs with transfer RNA (tRNA) in vitro and compare the results to those of the corresponding Pt-DNA complexes. Solutions containing various CD-37 or cisplatin concentrations were reacted with tRNA at physiological pH. Using Fourier transform infrared (FTIR), UV-visible, and circular dichroism spectroscopic methods, the drug binding mode, the binding constant, and RNA structural variations are determined for Pt-tRNA complexes in aqueous solution. Structural analysis showed direct binding of cisplatin drug to guanine and adenine N7 sites, while both direct and indirect interactions of CD-37 with tRNA bases and the backbone phosphate group were observed. The overall binding constants estimated were K(CD-37) = 2.77 (+/-0.90) x 10(4) M(1) and K(cisplatin) = 1.72 (+/-0.50) x 10(4) M(1). Major aggregation of tRNA occurs at high CD-37 concentrations, while RNA remains in the A-family structure.
Subject(s)
Antineoplastic Agents/metabolism , Cisplatin/metabolism , Estradiol/analogs & derivatives , Organoplatinum Compounds/metabolism , RNA, Transfer/metabolism , Binding Sites , Circular Dichroism , Estradiol/metabolism , Humans , In Vitro Techniques , Neoplasms/drug therapy , Neoplasms/metabolism , Nucleic Acid Conformation , RNA, Fungal/chemistry , RNA, Fungal/metabolism , RNA, Transfer/chemistry , Spectrophotometry, Ultraviolet , Spectroscopy, Fourier Transform InfraredABSTRACT
Platinum(II)-based anticancer drugs are effective for the management and treatment of several types of cancer. Cis-diamminedichloroplatinum(II) (cisplatin) exerts its antitumor activity by binding to DNA via intrastrand cross-links to d(GpG) (dG = deoxyguanosine) and to d(ApG) (dA = deoxyadenosine), causing DNA bending and interfering with DNA replication and transcription. However, the exact binding modes of other platinum(II)-based antitumor drugs to DNA duplex and their mechanism of action have not been clearly investigated. The aim of this study was to examine the binding of a novel anticancer estradiol-platinum(II) hybrid molecule (CD-37) with calf-thymus DNA in vitro and to compare the results with those obtained with cisplatin drug. Solutions containing various CD-37 or cisplatin concentrations were reacted with DNA at physiological pH. Then, using Fourier transform infrared, ultraviolet-visible, and circular dichroism spectroscopic methods, it was possible to characterize the drug binding mode, the binding constant, and structural variations of DNA in aqueous solution. Spectroscopic evidence showed that cisplatin binds to guanine N7 site with minor perturbations of the backbone phosphate group with an overall binding constant of K(cisPt) = 5.73 (+/- 0.45) x 10(4) M(-1). CD-37 binds to DNA duplex via H-bonding network at low drug concentrations with minor perturbations of guanine N7 site at high drug content and with a binding constant of K(CD-37) = 1.0 (+/- 0.15) x 10(4) M(-1). DNA aggregation occurs at high drug concentration, while DNA remains in the B-family structure.
