title: Is the contribution of cis and trans protonated 5-methylcytosine-SO3(-) isomers equal in the conversion to thymine-SO3(-) under bisulfite conditions? A theoretical perspective.
abstract: Cytosine (Cyt) can be converted to 5-methylcytosine (5-MeCyt) in CpG sequences of DNA. Conventional bisulfite sequencing can discriminate Cyt from 5-MeCyt, however inappropriate conversion of 5-MeCyt to thymine and a failure to convert Cyt to uracil always occur when Cyt and 5-MeCyt are treated with bisulfite, which would lead to erroneous estimates of DNA methylation densities. Here, the direct hydrolytic deamination of cis (paths A-C) and trans (paths A'-C') 5-MeCytN3(+)-SO3(-) isomers with bisulfite have been explored at the MP2/6-311++G(3df,3pd)//B3LYP/6-311++G(d,p) level. The activation free energies (ΔG(s-a≠)) of the cis and trans 5-MeCytN3(+)-SO3(-) isomers' paths exhibit no obvious differences, implying both isomers may make an equal contribution to the hydrolytic deamination of 5-MeCyt under bisulfite conditions. It is greatly expected that these results could aid experimental scientists to explore new methods to avoid the formation of the deaminated reactants (5-MeCytN3(+)-SO3(-)). Meanwhile, the HSO3(-)-induced direct hydrolytic deamination of cis and trans 5-MeCytN3(+)-SO3(-) isomers is represented by paths A and A', respectively, and has been further explored in the presence of two water molecules. It was found that the contribution of two water molecules renders the HSO3(-)-induced direct hydrolytic deamination of cis and trans 5-MeCytN3(+)-SO3(-) isomers by paths A and A' favourable. In addition, the ΔG(s-a≠) values (85.74-85.34 kJ mol(-1)) of the rate-limiting steps of the two water-mediated paths A and A' are very close to that of the theoretical value for CytN3(+)-SO3(-) (88.18 kJ mol(-1)), implying that the free barrier gap between Cyt and 5-MeCyt is very small under bisulfite conditions. This further suggests that bisulfite sequencing technology may be easily influenced by the external environment.
authors: Key Laboratory for Macromolecular Science of Shaanxi Province, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi'an 710062, China. wlwang@snnu.edu.cn.
authors: Lingxia
authors: L
authors: Jin
authors: Lu
authors: L
authors: Wang
authors: Caiying
authors: C
authors: Zhang
authors: Wenliang
authors: W
authors: Wang
authors: Suotian
authors: S
authors: Min
authors: Daodao
authors: D
authors: Hu
mesh_terms: D044503:5-Methylcytosine / Q000737:chemistry*; D007536:Isomerism; D008958:Models, Molecular; D011522:Protons; D013431:Sulfates / Q000737:chemistry*; D013447:Sulfites / Q000737:chemistry*; D013941:Thymine / Q000737:chemistry*
chemical_list: D011522:Protons; D013431:Sulfates; D013447:Sulfites; D044503:5-Methylcytosine; C042345:hydrogen sulfite; D013941:Thymine