Gas‐Phase Studies of Formamidopyrimidine Glycosylase (Fpg) Substrates

Abstract

Gas-phase thermochemical properties (tautomerism, acidity, and proton affinity) have been measured and calculated for a series of nucleobase derivatives that have not heretofore been examined under vacuum. The studied species are substrates for the enzyme formamidopyrimidine glycosylase (Fpg), which cleaves damaged nucleobases from DNA. The gas-phase results are compared and contrasted to solution-phase data, to afford insight into the Fpg mechanism. Calculations are also used to probe the energetics of various possible mechanisms and to predict isotope effects that could potentially allow for discrimination between different mechanisms. Specifically, (18) O substitution at the ribose O4' is predicted to result in a normal kinetic isotope effect (KIE) for a ring-opening "endocyclic" mechanism and an inverse KIE for a direct base excision "exocyclic" pathway.

Gas‐phase experimental and computational studies of human hypoxanthine‐guanine phosphoribosyltransferase substrates: Intrinsic properties and biological implications

“…As we noted earlier, in our experience, B3LYP/6‐31+G(d) yields accurate values for thermochemical properties of nucleobases. [ 25–36 ] For the purines studied herein, the calculated and experimental values in Table 3 are in agreement, within the experimental error, indicating that B3LYP/6‐31+G(d) appears to provide fairly accurate predictions for the thermochemical values. One exception is Apn PA, which we will discuss later in this section.…”

Gas-Phase Experimental and Computational Studies of 5-Halouracils: Intrinsic Properties and Biological Implications

“…Because the canonical tautomer is the most stable by about 10 kcal mol −1 for all four halouracils, we can assume that under our gas-phase conditions, the canonical structure is most likely the sole tautomer that is present. [3][4][5][6][7][8][9][10][11]15,18 Experimentally, there has been one prior measurement of 5-fluorouracil acidity by Chen and Chen (329 ± 4 kcal mol −1 ), which is consistent with our results. 32 Initially, we calculated 5FU, 5ClU, and 5BrU properties using B3LYP/ 6-31+G(d).…”

“…In our prior work with both normal and damaged nucleobases, we have found that B3LYP/6-31+G­(d) computational acidity and PA values generally compare quite favorably to experimental values. − ,, However, the 6-31+G­(d) basis set does not accommodate iodine, so we utilized the effective core potential def2-TZVP basis set of Ahlrichs and co-workers for the 5-halouracil calculations Figure shows the seven most stable calculated tautomers of 5FU ( 1 ); we show all tautomers that are within 15 kcal mol –1 of the canonical tautomer, 1a , as well as the acidity and PA of all the sites of all the tautomers.…”

“…Although the 6-31+G(d) basis set does not accommodate iodine (so we could not calculate the properties of 5IU), we were interested in the accuracy of this basis set for the other halouracils, as many of our prior calculations with other normal and damaged nucleobases had shown this basis set to be quite accurate. [3][4][5][6][7][8][9][10][11]15,18 As can be seen in Table 9, the B3LYP/6-31+G(d) calculations are not particularly consistent with the experimental data. For acidity, the trend between the B3LYP/6-31+G(d) calculations and experiment is the same (5FU is less acidic than 5ClU, which is less acidic than 5BrU), but the calculated values are 3−7 kcal mol −1 lower than the experimental data.…”

“…Damaged DNA bases differ in structure and properties from normal nucleobases and thus intervene with gene replication and expression, leading to carcinogenesis, cell death, and aging. , We have found that the contrast between the inherent properties of mutagenic, damaged bases, and normal bases lends insights into the toxicity, identification, and removal of damaged bases. − Our work has largely been motivated by understanding the processes that remove damaged bases from DNA. We have focused on the base excision repair (BER) pathway, particularly the mechanisms of glycosylase enzymes, which excise a wide range of mutated nucleobases. , Characterization of the intrinsic thermochemical properties lends insights into how damaged bases differ from normal bases, which in turn can reveal how mutations are selectively removed from the genome.…”