Detection of T-T Mismatches Using Mass Spectrometry: Specific Interactions of Hg(II) with Oligonucleotides Rich in Thymine (T)

Abstract

Electrospray ionization tandem mass spectrometry (ESI-MS/MS) was employed in a detailed study of the interactions of mercury dications with selected oligodeoxynucleotides rich and poor in thymine (T): d(5'-TT-3'), d(5'-TTT-3'), d(5'-TTTT-3'), d(5'-GG-3'), d(5'-CC-3'), d(5'-AA-3'), d(5'-GCTTGC-3'), d(5'-GTGCTC-3'), d(5'-GCATGC-3'), and d(5'-GCGCGC-3'). Specific interactions are observed for Hg(2+) with pure and mixed thymine sequences in which simultaneous bonding between two thymine units is indicated, and this is consistent with a model proposed in the literature in which Hg(2+) covalently coordinates to two thymines by replacing two N3 imino protons of the bases. The ESI-MS/MS measurements, combined with data on the thermal stability of mixed sequence hexadeoxynucleotides, indicate that mercury prefers thymines over the other binding sites in oligonucleotides both in solution and in the gas phase. These results point toward the effective use of Hg(2+) in the fast detection of mismatch base pairs incorporated in oligonucleotide duplexes and longer mixed DNA sequences using ESI-MS/MS.

Alkylation of uracil and thymine in the gas phase through interaction with alkylmercury compounds

“…It has been hypothesized that DNA double helix interactions with different mercury compounds (Hg 2+ , [RHg] + ) occur [45]. When interacting with DNA, mercury compounds also seem to exhibit a strong affinity for the thymine residues [46]. However, the detailed structure of adducts formed between [RHg] + compounds and DNA has yet to be determined.…”

Comparing ion‐pairing reagents and sample dissolution solvents for ion‐pairing reversed‐phase liquid chromatography/electrospray ionization mass spectrometry analysis of oligonucleotides

“…The emergence of soft ionization techniques for mass spectrometry, namely electrospray ionization (ESI) and matrix‐assisted laser desorption/ionization (MALDI), has substantially benefited the characterization of oligonucleotides by mass spectrometry (MS). However, mass measurement of oligonucleotides is complicated by the affinity of the polyanionic backbone to various cations including sodium and potassium which are often present in analyte solutions .…”

“…Therefore, quality control is a critical step in their synthesis and is usually accomplished by chromatographic techniques such as high-performance liquid chromatography (HPLC) [6][7][8][9][10] and electrophoresis. [11][12][13][14][15] The emergence of soft ionization techniques for mass spectrometry, namely electrospray ionization (ESI) and matrixassisted laser desorption/ionization (MALDI), [16][17][18][19][20][21] has substantially benefited the characterization of oligonucleotides by mass spectrometry (MS). However, mass measurement of oligonucleotides is complicated by the affinity of the polyanionic backbone to various cations including sodium and potassium which are often present in analyte solutions.…”

Analysis of oligonucleotides by ion-pairing hydrophilic interaction liquid chromatography/electrospray ionization mass spectrometry

“…Since Fenn and co‐workers demonstrated the use of ESI for the ionization of high‐mass biologically important compounds, electrospray ionization mass spectrometry (ESI‐MS) has shown its important advantages in the sequencing, identification, and characterization of oligonucleotides, as well as for applications in genotyping . However, mass measurement of oligonucleotides faces a serious problem of adduct formation, as the deprotonated phosphate backbone of oligonucleotides has a high affinity for various cations, in particular sodium and potassium ions, which are often present in analyte solutions .…”