Mass Spectrometry of Nucleic Acids

Abstract

Nucleic acids are prone to fragmentation in the ionization process of mass spectrometry (MS). The cause can be found in the polar nature of this analyte class. Following the introduction of the so‐called soft‐ionization techniques of electrospray ionization (ESI) and matrix‐assisted laser desorption ionization (MALDI) at the end of the 1980s, mass spectrometric analysis of nucleic acids has gained broader applicability. In combination with further developments of the associated instrumentation and optimized preparation and purification methods, a dramatic enhancement of the mass range, detection sensitivity, and resolution became possible in routine analysis of oligonucleotides, up to a length of 50 nucleotides. At present, the necessary quantity of sample is in the subfemtomole range. MS is an accurate, rapid, and sensitive tool ideally suited for sequencing shorter nucleic acids. MS sequencing holds great promise in clinical diagnostics, forensics, paternity analysis, livestock breeding, plant cultivation, and cell line typing. In addition, MS has been used for various other important applications including analysis of noncovalent complexes, mixture analysis, different sequencing strategies, misincorporation analysis of translesion DNA synthesis products, detection and mapping of DNA lesions, detection and characterization of DNA‐protein crosslinks (DPCs), and clinical diagnostics. The use of MS for the detection of DNA adducts in oligonucleotides is extremely useful because it has not only eliminated the need for hydrolysis but also allowed to map the position of the adduct and determine the sequence of the oligonucleotide. This article describes the fundamentals of mass spectrometric analyses of nucleic acids and some of its common applications. The primary focus is on LC‐ESI‐tandem MS and MALDI/MS.

Structural and Kinetic Analysis of Miscoding Opposite the DNA Adduct 1,N6-Ethenodeoxyadenosine by Human Translesion DNA Polymerase η

“…To gain insight into the ability of hpol to extend beyond the lesion, an LC-MS/MS method previously developed in this laboratory (34 -36) and applied extensively (37)(38)(39)(40)(41)(42)(43) was used for sequence analysis of the extension products. dU-containing primers were extended by hpol in the presence of all four dNTPs, followed by treatment with UDG and piperidine to cleave the fully extended products into shorter fragments for LC-MS/MS analysis (35,36).…”

A Review of Mass Spectrometry as an Important Analytical Tool for Structural Elucidation of Biological Products

“…Mass spectrometry based techniques is a rapid, accurate and sensitive tool ideally suited for structural elucidation of nucleic acids (Chowdhury & Guengerich, 2021). Mass spectrometry offers unique advantages due to its capability to distinguish each stoichiometry present in a mixture.…”