Methylcytosines

Primary synthesis has limited application in making pyrimidine-carboxylic acids or even their esters. However, some pyrimidine-4(and 5)-carboxylic acids can be effectively so made. For example, bromomucic acid (785) reacts as an aidehydo ketone with S-methyl-thiourea to give 5-bromo-2-methylthiopyrimidine-4-carboxylic acid (786) directly (53JCS3129) while the Whitehead synthesis (Section 2.13.3.1.2<7) can give, for instance, 3-methylcytosine-5-carboxylic acid (787) (55MI21300). 

Methylcytosine (964 X = O) was synthesized in 1901 and its isolation from hydrolyzates of tubercule bacilli was reported in 1925. However, this was later shown to be incorrect and only about 1950 was it isolated by hydrolysis of the deoxyribonucleotide fractions from thymus, wheat germ and other sources (50MI21302). Nucleotides and a nucleoside of 5-methylcytosine are known. 

Probably the best synthesis of 5-methylcytosine is via its 2-thio analogue (964 X = S) (49JBC(177)357) but the following briefly-described route may be preferable 2,4-dimethoxy-5-methylpyrimidine (965) with acetyl chloride gives the 1-acetyl derivative (966) which on aminolysis in aqueous ammonia gives 5-methylcytosine (964 X = O) in good overall yield (68TL2171). 

The Z-form can arise in sequences that are not strictly alternating Py-Pu. For example, the hexanucleotide " CGAT" CG, a Py-Pu-Pu-Py-Py-Pu sequence containing two 5-methylcytosines crystallizes as Z-DNA. Indeed, the in vivo... 

As is well-known, nucleic acids consist of a polymeric chain of monotonously reiterating molecules of phosphoric acid and a sugar. In ribonucleic acid, the sugar component is represented by n-ribose, in deoxyribonucleic acid by D-2-deoxyribose. To this chain pyrimidine and purine derivatives are bound at the sugar moieties, these derivatives being conventionally, even if inaccurately, termed as pyrimidine and purine bases. The bases in question are uracil (in ribonucleic acids) or thymine (in deoxyribonucleic acids), cytosine, adenine, guanine, in some cases 5-methylcytosine and 5-hydroxymethylcyto-sine. In addition to these, a number of the so-called odd bases occurring in small amounts in some ribonucleic acid fractions have been isolated. 

Todd and his associates - have established that iV-acetyl-1-methylcytosine exists predominantly as 270, rather than as 271, by... 

The reaction mechanism of the DNA (cytosine-5)-methyltransferase-catalyzed cytosine methylation was investigated at the MP2 and DFT levels [98JA12895]. This system has been modeled by 1-methylcytosine 117, methylthiolate, and trimethylsulfonium. The cytosine methylation is initiated by an attack of the anionic methylthiolate at Cg of the cytosine ring (Scheme 77). The formation of the methylthiolate adduct 118 of the neutral 117 was found to be endothermic in the gas phase and in solution. However, the MP2 and DFT results differ... 

Among the simplest compounds of this series are l-(2-deoxy-2-fluoro- -D-arabinofuranosyl)thymine (758, FMAU) and 1-(2-deoxy-2-fluoro-)S-D-arabinofuranosyl)-5-methylcytosine (749, FMAC). Later FIAU (757) and FMAU (758) were prepared in high yields (>95% the ratio of Plot an-omers being 7/1 and 4/1, respectively) by condensation using 743 and 2,4-bis(0-trimethylsilyl)-5-iodouracil (769), and 743 and 2,4-bis(0-trimethylsi-lyl)thymine (770), respectively. Likewise, 753, 757, 758, and 765 (this was described later) were prepared in high yields using pyrimidine derivatives... 

Small quantities of additional purines and pyrimidines occur in DNA and RNAs. Examples include 5-methyl-cytosine of bacterial and human DNA, 5-hydroxy-methylcytosine of bacterial and viral nucleic acids, and mono- and di-N-methylated adenine and guanine of... 

Nucleic acids contain, in addition to A, G, C, T, and U, traces of 5-methylcytosine, 5-hydroxymethylcyto-sine, pseudouridine ( P), or N-methylated bases. 

Fig. 25. Heterometallic complex of 1-methylcytosine, highlighting the nature of the metal-metal interaction involving the filled Ptn dz2 with the Pd11 dl2 v2.

The data in Table 2.1 suggest that the O-benzylated adduct cannot be isolated since it is less stable than reactants. The N3-benzylated adduct should be generated faster, but it should also decompose under mild conditions into free reactants, because the activation free energy in aqueous solution for the decomposition into free QM and methylcytosine is only 21.4 kcal/mol.14 In other words, these data suggested that the QM-N3-cytosine conjugate could act as QM-carrier, few years before the experimental data related to the stability of QM-conjugates became available.4... 

One-Electron Oxidation Reactions of Cytosine and 5-methylcytosine DNA Base... 

In 1 JMSO-z4, or DMF-(/, the 1 1 condensation products of cytosine or iV-methylcytosine with triformylmethane, compounds 39a and 39b, showed ring-chain tautomerism with the ring-closed products 40a and 40b (see Equation (2), Section 12.04.2.4) <1996ACS1137>. 

Not all mutagenesis in IS. coli is dependent on SOS-processing. Mutations may arise quite simply during DNA replication if a base is substituted by or converted to another, incorrect, base. Consider the consequence of oxidative deamination of the base 5-methylcytosine to thymine. Replication followed by daughter strand segregation will result in a G C base pair having been mutated to an A T base pair. Sites containing 5-methylcytosine are hotspots for G C to A T transitions in 12. coli (24). 

Sites containing 5-methylcytosines (CCAGG). These are spontaneous lacl hotspots. 

Figure 1. The frequencies of amber mutations in the lacl gene induced by BPDE. Solid bars, individual sites at which we detected mutations open bars, sites at which we did not detect mutations asterisks (in a), sites at which the target codon contains 5-methylcytosine.

For various reasons, the generalizations mentioned above must be regarded as strictly provisional. Analyses utilizing formic acid indicate the presence of more than one phosphorus atom per purine or pyrimidine residue. This discrepancy, it is pointed out, could equally well result from an apparent deficiency of bases, due to error in the analytical technique.160 It is also necessary to consider that some nucleic acids are now known to contain more bases than was previously realized. Thus, 5-(hydroxymethyl)-cytosine is present in various viruses,181-182 and 5-methylcytosine occurs in various animal and plant deoxyribonucleic acids but is absent from those of microbial origin.17-160-1M- 184- 186 Certain microbial deoxyribonucleic acids also contain 6-methylaminopurine.186a Various bacteriophage deoxyribonucleic acids have been found to contain a component which is believed to consist of a D-glucoside186b of 5 -(hydroxymethyl)cytidylic acid. 

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