7-methylguanosine and 1-methylguanosine

Laser ionization of guanosines containing methyl substitutions in the 1-, N2-, 3'-O-, O6- and 7-positions generated two characteristic negative ions: loss of hydrogen to generate [M - H]- and elimination of the sugar ring to form the nucleic base ion. The ions generated by elimination of the sugar ring provided the information necessary to determine whether the methyl group was on the nucleic base or sugar ring. Fourier transform mass spectrometry was used to isolate and collisionally dissociate selected negative ions from these nucleosides. The collisional dissociation spectra indicated daughter ions which were sufficient to differentiate all the isomers with methyl substitution of the nucleic bases. In addition, accurate mass measurement and sequential collisional dissociation experiments were employed to investigate fragmentation mechanisms.

Topics: Guanosine; Mass Spectrometry

Mitomycin C (1) is a clinically used antitumor antibiotic that binds covalently to deoxyribonucleic acid under reductive or acidic catalysis. We have determined the structures of the adducts resulting from attack of reductively activated 1 on the dinucleoside phosphate d(GpC) to be N2-(2'' beta, 7''-diaminomitosen-1''alpha-yl)-2'-deoxyguanosine (2) and its 1'' beta-isomer (3). This represents a revision of the previously reported structures for these adducts in that the mitomycin residue is linked to the N2- rather than O6-position of 2'-deoxyguanosine. This revision is the result of applying to the mitomycin case a newly developed general method that leads to unambiguous assignment of the linkage position in complex alkylated guanosines. The method as described here takes advantage of the resolution enhancement gained by calculation of the second derivatives of absorbance Fourier transform infrared spectra. In addition, we present 1H NMR data that corroborate the assigned structures of 2 and 3 and that should serve as a useful reference for future investigations into the binding of mitomycin C to DNA. The convenient synthesis of adducts 2 and 3 from deoxyguanosine and mitomycin C reported here should facilitate such investigations as well. Furthermore, we demonstrate a useful acetylation procedure for adducts and metabolites of mitomycin C that furnishes spectroscopically superior chemical derivatives (e.g., triacetates 4 and 5, derived from acetylation of adducts 2 and 3).

Topics: Deoxyguanosine; Fourier Analysis; Guanosine; Magnetic Resonance Spectroscopy; Mitomycin; Mitomycins; Spectrophotometry, Infrared

A fast method for a single-step fractionation of a number of tRNA methyltransferases from Salmonella typhimurium is described. The method basically consists of ion-exchange chromatography on a phosphocellulose column and permits the separation of the enzymes forming mt6A, m1G, m5U, m7G. The enzyme fractions appear sufficiently purified to allow the estimation of some molecular and kinetic properties. The apparent KM for adenosylmethionine range between 1.5 to 3.2 X 10(-5) M, whereas KM for undermethylated tRNA range between 3.1 X 10(-5) M to 3.1 X 10(-4) M. Glycerol gradient determination indicates the following Mr for the native proteins: 25 X 10(3), 40 X 10(3), 50 X 10(3) and 65 X 10(3) for m7G-, mt6A-, m1G- and m5U-forming enzymes, respectively. A complete analysis of methylated nucleosides formed in vivo in S. typhimurium has been obtained: it also allowed us to infer the pattern of the various tRNA methyltransferases for this prokaryote. The tRNA methyltransferase forming mt6A has been isolated for the first time from any type of cell.

Topics: Chromatography, Ion Exchange; Chromatography, Thin Layer; Guanosine; Kinetics; Molecular Weight; Purine Nucleosides; Salmonella typhimurium; Threonine; tRNA Methyltransferases; Uridine

The binding interaction of aquated cis-(NH3)2Pt(II) with 1-methylguanosine (1-MeGuo), 7-methylguanosine (7-MeGuo), 1-methyladenosine (1-MeAdo+) and protonated adenosine has been studied using UV difference spectroscopy. The magnitude of the binding constants for the 1 : 1 interaction of cis-(NH3)2Pt(II) with 1-MeDuo and 1-MeAdo are log K = 3.5 and 3.7, respectively. These data are presented and compared to other cis-(NH3)2Pt(II)-nucleoside interactions.

Topics: Adenosine; Binding Sites; Cisplatin; DNA; Guanosine; Nucleosides; Spectrophotometry, Ultraviolet