Transalkylation of Phosphotriesters Using Cob(I)alamin:  Toward Specific Determination of DNA−Phosphate Adducts

Abstract

The supernucleophilic cobalt compound, cob(I)alamin, has been kinetically characterized with respect to its ability to bring about transalkylation of adducts to DNA phosphates (phosphotriesters). The reactivity of cob(I)alamin toward different phosphotriesters (model compounds and methylated DNA), as well as its specificity toward DNA-phosphate adducts, has been investigated. Through nucleophilic displacement on the alkyl by cob(I)alamin, the alkyl groups (methyl and ethyl) were transferred from phosphotriesters within minutes at room temperature. In contrast, methylated nucleosides (base adducts) were stable in the presence of cob(I)alamin.

Kinetic and Mechanistic Studies on the Reactions of the Reduced Vitamin B12 Complex Cob(I)alamin with Nitrite and Nitrate

“…[4] Cbl(I) dehalogenates or dealkylates a wide range of organic molecules, including alkanes, alkenes, alkynes, aromatics, and phosphotriesters. [4,11] There is also much interest in using Cbl(I) to trap reactive intermediates such as oxiranes. [12] Kinetic studies have been reported for the dehalogenation of halogenated alkanes, [13] alkenes, [14] and acetic acid [15] and the dealkylation of phosphotriesters.…”

Analysis of DNA-phosphate adducts in vitro using miniaturized LC-ESI-MS/MS and column switching: Phosphotriesters and alkyl cobalamins

“…The hydrolysate of ENU-treated DNA was transalkylated using cob(I)alamin according to an earlier developed method [17]. DNA sample (100 L) was added to a reduced cobalamin solution under argon.…”

“…The program used was the following: Samples were injected on the pre-column using the auxiliary system C at [15][16][17][18][19][20] L/min. At the same time the binary system A/B pumps 20% Solvent B at a flow of 2-8 L/min over the analytical column.…”

“…For comparison, identical DNA hydrolysate was analyzed using an independent method, the so-called transalkylation method. This method comprises of specific transfer of DNA-phosphate adducts to the nucleophilic cob(I)alamin [17,18]. The resulting ethyl cobalamins (EtCbl) were analyzed according to an earlier report [19].…”

Characterization of alkyl‐cobalamins formed on trapping of epoxide metabolites of 1,3‐butadiene

“…The usefulness of Cbl as an analytical tool has been demonstrated in earlier studies where Cbl has been utilized for transfer and analysis of adducts to phosphate oxygens in DNA [7] and for investigation of the reactivity of 1,2-epoxides [8]. The main advantage of Cbl compared to other standard nucleophiles, e. g., thiosulfate and nicotinamide [7,8] is that Cbl in its reduced form [Cbl(I)] is by orders of magnitude more reactive.…”