Selective Cleavage of a 35‐mer Single‐Stranded DNA Containing the Initiation Codon of the TAT Gene of HIV‐1 by a Tailored Cationic Manganese Porphyrin

Abstract

volume of the loaded VPI-5 was only about one third of that of an unloaded sample (unloaded: 0.184 ~m -~, loaded 0.065 cm3).Besides the proof of the inclusion of C,, in the micropores, this study also included preliminary investigations on the dynamics of the adsorbed C60. For this purpose solid-state I3C N M R spectra both with and without rotation about the magic angle (magic angle spinning, MAS) were recorded for aluminophosphate samples that have no detectable bulk C,, on the outer surface according to XRD. These spectra showed that the C,, molecules rotate freely within the channels of the aluminophosphates, because the line-broadening expected for restricted rotationf51 was not unambiguously observed. Nevertheless, the lines were broadened in those spectra recorded without MAS, from which a partial restriction of the free isotropic rotation can be deduced. A further, though only slight, broadening of the lines from 600 to 700 Hz was obtained by cooling the sample from 298 to 230 K. This might point to a transition from the "rotator" phase to the "rachet" phase (the low-temperature phase according to ref. (51) in the aluminophosphate lattice at relatively high temperatures.['] It should, however, be mentioned that coadsorbed water can influence the dynamics significantly.At present we are studying in more detail the electronic and optical properties of the molcular sieves doped with C,, .In particular, we are attempting to separate C,,, C,,, and higher fullerenes in the gas phase by these methods. This would be an alternative to the usual separation by liquid chromatography['. and is also applicable on larger scales. E.xperinrenta1 ProcedureSample preparation: AIPO,-5 and AIPO,-8 were activated by simple heating to 200 C in air to remove adsorbed water. Under analogous treatment VPI-5 was transformed irreversibly into AIPO,-X and had therefore to be dehydrated under high vacuum Torr) [4]. The aluminophosphate samples were placed in one arm of an inverted glass U-tube. and the other was charged with C60. The arm containing aluminophosphate was heated separately. and the sample chamber was evacuated prior to calcination through the opening in the center of the tube. At lo-' Torr the aluminophosphates were first activated at 280 "C for 12 hand then at 480 "C for 1 h. Thereafter under maintenance of the vacuum the glass tube was fused, and the glass ampule containing the activated aluminophosphate and C,, was heated for 10 h at 450 C. The tube was opened, and the sample washed in toluene. filtered off'. and again washed for three periods of 15 min in toluene (200 mL portions per 1 g loaded aluminophosphate) in a ultrasonic bath. This procedure removed the last residues of C,, from the crystal surface (determined by XRD). The IR spectra were recorded on crushed crystal aggregates with an Analytical Plan IR microscope (SpectraTec) coupled to a Nicolet 5SXB FTIR spectrometer. The N L sorption measurements were performed at 77 K with a volumetric sorption apparatus (Omnisorp 100). To remove adsorbed water and toluen...

Preparation of hybrid "DNA cleaver-oligodeoxyribonulceotide" molecules based on a metallotris (methylpyridiniumyl)porphyrin motif

“…Preliminary experiments (21) indicate that the vectorized cationic manganese porphyrin 8 is able to recognize ita complementary sequence incorporated in a 35-mer, even in the presence of a large excess of salmon sperm DNA, and to cleave the target at very low concentrations (5-50 nM) when activated by potassium monopersulfate, an oxygen atom donor that we used to mimic bleomycin activity (15, 22). Work is in progress to develop these vectorized cationic manganese porphyrins having an efficient nuclease activity with a low cleaverbarget ratio (1 to 5/11 as antiviral agents or antioncogens.…”

DNA as Platform for New Biomaterials. Metal-Containing Nucleic Acids

“…Among different porphyrin motifs one of the most effective appeared the diaquamanganese (III) meso-tetrakis(N-methylpyridinium-4-yl)porphyrin [99]. The first generation of metalloporphyrin/DNA conjugates consisted of complexes with the metalloporphyrin moiety attached to the 5'-end of the DNA-oligonucleotide.…”

Oxidative damage on RNA mediated by cationic metalloporphyrin–antisense oligonucleotides conjugates †

“…Antisense 5Ј-OH oligonucleotides or antisense metalloporphyrin-ODN conjugates were complementary to the region (6,9), 500 n (7, 10) or 1 µ (8, 11) of AS-2-Mn in the presence of RNase H (6-8) or KHSO 5 (9)(10)(11); 12-17, RNA incubated with 100 n (12,15), 500 n (13,16) or 1 µ (14, 17) of S-2-Mn in the presence of RNase H (12)(13)(14) or KHSO 5 (15)(16)(17); 18-20, free metalloporphyrin assayed at 100 n (18), 500 n (19) or 1 µ (20) in the presence of KHSO 5 (ϩ79,ϩ96) of the CAT sequence containing the translation initiation codon of the CAT protein. They were incubated with the non-labelled 838-base RNA substrate in the presence of KHSO 5 or RNase H and the resulting non-labelled hybrids of damaged RNA/antisense were then taken as template for the reverse transcription reaction performed with [α-32 P]dCTP.…”