Formation of Aminyl Radicals on Electron Attachment to AZT: Abstraction from the Sugar Phosphate Backbone versus One-Electron Oxidation of Guanine

Abstract

Employing electron spin resonance (ESR) spectroscopy, we have characterized the radicals formed in 3′-Azido-3′-deoxythymidine (3′-AZT) and in its 5′-analog 5′-azido-5′-deoxythymidine (5′-AZT) after electron attachment in γ-irradiated aqueous (H 2 O or D 2 O) glassy (7.5 M LiCl) systems. ESR spectral studies and theoretical calculations show that the predominant site of electron capture in 3′-AZT and in 5′-AZT is at the azide group and not at the thymine moiety. The azide group in AZT is therefore more electron affinic than the most electron affinic DNA base, thymine. Electron attachment to 3′-AZT and 5′-AZT results in an unstable azide anion radical intermediate (RN 3 • − ) that is too short lived to be observed in our work even at 77 K. At 77 K we observe the neutral aminyl radical (RNH•) after loss of N 2 from RN 3 • − followed by protonation of nitrene anion radical (RN• − ) to give RNH•. The expected RN• − intermediate is not observed as protonation from water is complete at 77 K even in under highly basic conditions. Formation of RND• in D 2 O solutions confirms water as the source of the NH proton in the RNH•. Our assignments to these radicals are aided by DFT calculations for hyperfine coupling constants which closely match the experimental values. On annealing to higher temperatures (ca. 160-170 K), RNH• undergoes bimolecular hydrogen abstraction reactions from the thymine methyl group and the sugar moiety resulting in the formation of the thymine allyl radical (UCH 2 •) and two sugar radicals -C3′•, C5′•. RNH• also results in one-electron oxidation of the guanine base in 3′-AZG. This work provides a potential mechanism for the reported radiosensitization effects of AZT.

Insight Into the Inhibition of Ribonucleotide Reductases by 2'-chloro-2'-deoxynucleotides and 2'-azido-2'-deoxynucleotides: Biomimetic Studies with Model Substrates

“…79 Azide group in the modified nucleoside showed more attraction towards the electron compared to the thymine moiety which is consistent with the theoretical calculations and the ESR spectral calculations.…”

“…Based on Sevilla's work on 3′-azido-3′-deoxythymidine (3′-AZT) and 5′-azido-5′-deoxythymidine (5'-AZT) analogues, 79 …”

“…Following the work regarding the formation of radicals in 3'-AZT and the 5′-azido-5′-deoxythymidine (5'-AZT) analogues as well as 2'-azido-2'-deoxy uridine after attachment of electron in γ-irradiated aqueous glassy (7.5 M LiCl) systems, 79 I planned to extend my efforts to study radiation-generated electron-induced site specific formation of neutral aminyl radicals (RNH•) and their reactions in 2'-azido-2'-deoxycytidine (2'-AZdC,…”

“…Following our works, 79,141,142 these homogenous solutions were thoroughly bubbled with nitrogen gas and then immediately were drawn into 4 mm Suprasil quartz tubes (cat. No 734-PQ-8 WILMAD Glass co., Inc., Buena, NJ).…”

“…Following our works, 79,141,142 these glassy samples were γ-irradiated with the aid of 109-GR a irradiator containing shielded 60 Co source with an absorbed dose 525 -700…”