Glutathione-catalyzed hydrogen isotope exchange at position 5 of uridine. Model for enzymic carbon alkylation reactions of pyrimidines

Abstract

H-isotope exchange inhibitor are reasonably high as early as 12 hr after birth and increase dramatically as the animal becomes an adult. The data do not eliminate the birth process itself as being the triggering mechanism, however, studies carried out with fasted animals show that it is possible to delay the appearance of inhibitor activity by withholding food from the newborn animal for at least 48 hr.Recently, Sharma and Borek (1970) have shown a tRNA methylase inhibitor to be under estradiol regulation in both rat and pig uteri. In pigs, growth hormone levels are high at birth and remain elevated in animals that are starved for 60 hr after birth. In contrast, growth hormone levels decrease in animals fed immediately after birth to those levels observed in adult pigs (Swiatek et al., 1968). It would be interesting to determine whether the differences in levels of inhibitor in fed and fasted newborn pig brain are hormonally regulated or dependent on some other dietary factors as yet unexplained. Experiments are currently in progress to answer this question.

The Synthesis of Deuterionucleosides

“…Uridine-5-2 H can be isolated in good yield with high isotope incorporation upon treatment with 2-mercaptoethylamine in 2 H 2 O either at room or at elevated temperature 64,65 . Similar catalytic activity has been observed for glutathione 57 and cysteine 56,61 59 upon formation of a C6 adduct with 2'-deoxyuridine monophosphate.…”

“…This makes the NMR spectral assignment easier for structural studies [44][45][46][47][48][49][50][51] . Base deuterated molecules can also be used in conformational investigations by Raman [52][53][54] or infrared 55 spectroscopy, in biological studies [56][57][58][59][60][61] and studies on hydroxyl radical reactions with nucleosides 62,63 .…”

Amino acid sequence at the FdUMP binding site of thymidylate synthetase.

“…Although model studies favoring a nucleophilic attack at the 6-position of the pyrimidine ring have been reported (7,8), the role of cysteine as a nucleophile (9) was questioned recently by Sommer and Santi (10) when they could not obtain evidence for cysteine in an FdUMP-containing peptide isolated from the synthetase. The latter finding can be questioned, in turn, as a result of prior studies with sulfhydryl inhibitors (3,5,6), which implicated a sulfhydryl group at the active site and more recent ones with a5S-labeled thymidylate synthetase (11).…”

Enzyme inhibition as a source of new drugs

“…Thymidylate synthetase, a target of drugs in cancer chemotherapy [Danen- berg, 1977;Friedkin, 1973;Santi, 19801 is a good example for the operation of the principle of microscopic reversibility . Figure 9 outlines the mechanism of covalent nucleophilic catalysis of C-C bond formation, the initial sequence of a complex, multistep reaction [Benkovic and Bullard, 1973;Benkovic, 1980;Friedkin, 1973;Kalman, 1971a;Kalman, 197 lb;Pogolotti and Santi, 19771. It is also shown that an analogous reaction sequence is involved when the substrate, deoxyuridylate (dUMP) is replaced by the potent inhibitor, 5-fluoro-dUMP, the active form of the antineoplastic drugs, 5-fluoropyrimidines.…”