Mass Spectrometry and Proteomics 2018: the Mass Spectrometry Society of Japan,Japanese Proteomics Society,and Asia-Oceania Human Proteome Organization

ABSTRACT

Introduction: The mass spectrometry society of Japan, Japanese proteomics society, and Asia–Oceania human proteome organization held the conference ‘Mass Spectrometry and Proteomics 2018’ in Osaka, Japan, on May 15–18, 2018. This international conference focused on cutting edge technologies and their applications in a variety of research fields such as agriculture, material science, environmental factors, and clinical applications. An overview of the conference and a summary of the major lectures are reported here.

Expert commentary: The meeting will facilitate the development of fundamental technologies and the multi-disciplinary applications of proteomics.     

Recent Developments in Mass Spectrometry: Rna Sequencing

Abstract

Simplified sequencing of an oligoribonucleotide containing 16 bases is accomplished by matrix assisted laser desorptiodionization time of flight (MALDI-TOF) mass spectrometry. We used delayed ion extraction (DE) technique and kinetic degradation with two exonucleases.     

N-Dimethylaminomethylene-O-trialkylsilyl Derivatives of Nucleosides for Chromatography and Mass Spectrometry

Abstract

Dimethylformamide dimethyl acetal (DMF-DMA) reacts with nucleosides under mild conditions to give N-dimethylaminomethylene (N-DMAM) derivatives. Silylation provides the DMAM-O-trialkylsilyl mixed derivatives which have good chromatographic and mass spectral properties.     

Fast atom Bombardment Mass Spectrometry of Nucleosides,Nucleotides and Oligonucleotides

Abstract

Fast atom bombardment (FAB) mass spectrometry, a new ionization technique, has been applied to a variety of polar, nonvolatile compounds with considerable success. Current literature regarding the analysis of nucleosides, nucleotides and oligonucleotides using FAB is reviewed.     

Detection of Biosignatures by Geomatrix-Assisted Laser Desorption/Ionization (GALDI) Mass Spectrometry

Identification of mineral-associated biosignatures is of significance for retrieving biochemical information from geological records here on Earth and for detecting signs of life on other planets, such as Mars. An investigation using laser desorption Fourier transform mass spectrometry was conducted to determine whether geomatrix-assisted laser desorption/ionization (GALDI) can be used to detect amino acids (e.g., histidine, threonine, and cysteine) and small proteins (e.g., gramicidin S) associated with mineral phases and whether the geomatrix impacts detection. Iron oxide (Fe₂ O ₃ ) and sodium chloride (NaCl) were investigated as clean chemical analogues of hematite and halite, respectively, which have both been detected on the surface of Mars. Samples were prepared by 2 methods: (1) application of analyte solution to the geomatrix surface with subsequent drying; and (2) physical mixing of analyte and geomatrix. Amino acids incorporated within NaCl by physical mixing yielded a better signal-to-noise ratio than those that were applied to the surface of a NaCl pellet. The composition of the geomatrix had an influence on the detection of biomolecules. Peaks corresponding to the cation-attached biomolecular ions were observed for the NaCl prepared samples. However, no biomolecular ion species were observed in samples using Fe ₂ O ₃ as geomatrix. Instead, only minor peaks that may correspond to ions derived from fragments of the biomolecules were obtained.     

Rapid Diagnosis of Lung Tumors,a Feasability Study Using Maldi-Tof Mass Spectrometry

ObjectiveDespite recent advances in imaging and core or endoscopic biopsies, a percentage of patients have a major lung resection without diagnosis. We aimed to assess the feasibility of a rapid tissue preparation/analysis to discriminate cancerous from non-cancerous lung tissue.MethodsFresh sample preparations were analyzed with the Microflex LT^TM MALDI-TOF analyzer. Each main reference spectra (MSP) was consecutively included in a database. After definitive pathological diagnosis, each MSP was labeled as either cancerous or non-cancerous (normal, inflammatory, infectious nodules). A strategy was constructed based on the number of concordant responses of a mass spectrometry scoring algorithm. A 3-step evaluation included an internal and blind validation of a preliminary database (n = 182 reference spectra from the 100 first patients), followed by validation on a whole cohort database (n = 300 reference spectra from 159 patients). Diagnostic performance indicators were calculated.Results127 cancerous and 173 non-cancerous samples (144 peripheral biopsies and 29 inflammatory or infectious lesions) were processed within 30 minutes after biopsy sampling. At the most discriminatory level, the samples were correctly classified with a sensitivity, specificity and global accuracy of 92.1%, 97.1% and 95%, respectively.ConclusionsThe feasibility of rapid MALDI-TOF analysis, coupled with a very simple lung preparation procedure, appears promising and should be tested in several surgical settings where rapid on-site evaluation of abnormal tissue is required. In the operating room, it appears promising in case of tumors with an uncertain preoperative diagnosis and should be tested as a complementary approach to frozen-biopsy analysis.     

New Techniques for the Rapid Characterization of Oligonucleotides by Mass Spectrometry

Abstract

Recent advances in combined HPLC/electrospray ionization-mass spectrometry provide effective new capabilities for the rapid characterization of oligonucleotides. Accurate mass measurements with errors <0.3 Da, and determination of base and sugar modification and of nearest neighbor identities, can be routinely carried out on 10-100 component mixtures of RNA or DNA. These procedures are widely applicable in structural and analytical studies involving mixtures of oligonucleotides.     

Differentiation of Isomeric Purine and Pyrimidine Mononucleotides by Fast Atom Bombardment Tandem Mass Spectrometry

Abstract

The use of positive ion fast atom bombardment mass-analysed ion kinetic energy (FAB/MIKE) spectroscopy to differentiate the 2′, 3′-and 5′-monophosphate isomers of adenosine, guanosine and cytidine is described.     

Analysis of Substituted Pyridine-C-Nucleosides by Direct Liquid Introduction Liquid Chromatography/Mass Spectrometry

Abstract

A method was elaborated for the analysis of a few original pyridine-C-nucleosides via microbore DLI/LC-MS. The compounds were analyzed on a 10RP8 column (25 cm × 1 mm) using a number of 0.01 M HCOONH₄/CH₃OH mixtures as eluant. Under appropriate LC-MS conditions, both α- and β-anomers were separated and identified. All nucleosides were characterized by the protonated molecular ion [MH]+, [B+30]+ and [B+44]+-fragment ions. Assignment of the α, β-configuration at C₁′ was done with the aid of ¹³C-NMR. From the DLI/LC-MS data, a semi-preparative HPLC-method was developed to purify the pyridine-C-nucleosides prior to biological evaluation.     

Dna Analysis by Mass Spectrometry: Applications in Dna Sequencing And Dna Diagnostics

Abstract

Use of mass spectrometry to detect PCR amplified DNA from individuals infected with hepatitis B virus, to derive DNA sequence information through exo-nuclease based degradation or Sanger sequencing and a new format (PROBE) for the efficient determination of mutations is described.     

Time-of-flight Secondary Ion Mass Spectrometry of Branched RNA Fragrants: Messenger RNA Splicing Intermedistes

Abstract

The negative ion mass spectra generated by a reflecting time-of-flight mass spectrometer are reported for a series of protected oligonucleotides. Quasimolecular and sequence ions have been detected, and the location and nature of protecting groups have been confirmed.     

9th Annual Symposium on Advances in Separation Science and Mass Spectrometry

Evaluation of: Di Girolamo F, Boschetti E, Chung MC, Guadagni F, Righetti PG. ‘Proteomineering’ or not? The debate on biomarker discovery in sera continues. J. Proteomics 74(5), 589–594 (2011).

The combinatorial peptide ligand library in association with mass spectrometry can greatly enhance the dynamic range of the analysis of low- and very low-abundance proteins constituting the vast majority of species in any sample. When compared with untreated samples, the increment in detection of low-abundance species appears to be at least fourfold. Recently, the combinatorial peptide ligand library has been challenged; however, it has been clearly demonstrated in the evaluated paper that the protocols for elution of the captured polypeptides make the difference. Therefore, the solid-phase ligand library made of hexapeptides remains a promising and unique tool for biomarker discovery.     

Pancreatic Cancer Surgical Resection Margins: Molecular Assessment by Mass Spectrometry Imaging

BackgroundSurgical resection with microscopically negative margins remains the main curative option for pancreatic cancer; however, in practice intraoperative delineation of resection margins is challenging. Ambient mass spectrometry imaging has emerged as a powerful technique for chemical imaging and real-time diagnosis of tissue samples. We applied an approach combining desorption electrospray ionization mass spectrometry imaging (DESI-MSI) with the least absolute shrinkage and selection operator (Lasso) statistical method to diagnose pancreatic tissue sections and prospectively evaluate surgical resection margins from pancreatic cancer surgery.ConclusionsOur findings provide evidence that the molecular information obtained by DESI-MSI/Lasso from pancreatic tissue samples has the potential to transform the evaluation of surgical specimens. With further development, we believe the described methodology could be routinely used for intraoperative surgical margin assessment of pancreatic cancer.     

Mass Spectrometry: Applications in the Analysis of Nucleosides,Nucleotides and Oligonucleotides

Abstract

The past ten years have been an exciting time in mass spectrometry as a number of important instrumental developments have revolutionized the field, including the analysis of nucleic acid components.^1,2 The focus of this talk will be on the impact that new ionization methods, e.g., plasma desorption(PD) and fast atom bombardment(FAB), and new magnet technology (expanded mass range and scan speed capability) have had on the analysis of nucleosides and nucleotides. Results from the speaker's laboratory will be used to illustrate the significance of capillary GC/MS techniques for the separation and analysis of complex mixtures of nucleosides derived from a biological source. In addition, some approaches being developed to overcome current limitations in the FAB analysis of nucleosides and nucleotides will be described. Unfortunately, time does not permit a discussion of other new areas of interest, i.e., LC/MS³ and MS/MS.⁴     

Peptide Sequencing by Mass Spectrometry for Homology Searches and Cloning of Genes

It is now possible to obtain sequence information from gel-separated proteins by mass spectrometry at levels too low for conventional approaches. Usually this tandem mass spectrometric data are used for database searches with the aim of identifying the corresponding gene. Recently it has been shown that long and accurate amino acid sequences can be obtained which are sufficient for PCR-based strategies to clone the corresponding gene [Wilm et al. (1996), Nature 379, 466–469]. More than eight proteins have now been cloned based on that method. In many more cases the sequence information identified homologous proteins. Issues involved in cloning by mass spectrometric sequence information are discussed, as are two case studies. These results clearly establish mass spectrometry as a viable tool not only for the database identification of proteins, but also for the de novo sequencing of gel-separated proteins at the low-picomole to femtomole level.     

Synthesis and Mass Spectrometry Analysis of Oligonucleotides Bearing 5-Formyl-2′-Deoxyurldlne in Their Structure

Abstract

Two oligonucleotides containing FdU (1) have been synthesized. The use of the “Pac-amidites” for the natural nucleosides has allowed the incorporation of the oxidized thymine residue without protection of the aldehydic function. The oligonucleotide composition was confirmed by enzymatic digestion and electrospray mass spectrometry.     

MALDI-TOF Mass Spectrometry as a Powerful Tool to Study Enzymatic Processing of DNA Lesions Inserted into Oligonucleotides

Abstract

MALDI-TOF mass spectrometry measurements, coupled with either exonuclease or DNA N-glycosylases digestions of lesion-containing oligonucleotides, were used to assess biochemical features of several oxidative DNA damage. The latter analytical approach was shown to be an informative and efficient alternative technique to conventional electrophoresis and chromatographic analyses.     

Evaluation of Capillary HPLC/Mass Spectrometry as an Alternative Analysis Method for Gel Electrophoresis of Oligonucleotides

Abstract

A method has been developed to monitor the enzymatic incorporation of nucleotides in DNA by electrospray HPLC mass spectrometry. The main advantages of mass spectrometry over electrophoresis are the ability to directly characterize the reaction products and the shorter analysis time.     

Analysis of Antiretroviral Nucleosides by Electrospray Ionization Mass Spectrometry and Collision Induced Dissociation

Abstract

Antiretroviral nucleoside drugs used against the human immunodeficiency virus (HIV) infection have been analyzed using negative ion electrospray ionization (ESI) mass spectrometry and collision-induced dissociation (CID-MS/MS). Mass fragmentation of azidothymidine (AZT), didanosine (ddI), dideoxycytidine (ddC) and dideoxythiacytidine (3TC) were obtained at different cone voltages and collision energies. Fragmentation of purines and pyrimidines occurred by different pathways. For purines (ddI), the fragmentation was similar to those found in endogenous nucleosides; mainly the pseudo molecular ion is present (M-H) and a cleavage through the glycosidic bond forming (B) was observed. For pyrimidines (AZT, ddC, 3TC), the fragmentation pathways were different from endogenous nucleosides; for AZT, the fragmentation occurred primarily through the elimination of the azido group in the 3′-position (M-H₂-N₃), whereas ddC and 3TC presented more complex fragmentation patterns. For ddC, fragmentation appeared to be dominated by a retro Diels-Alder mechanism (M-CONH). For 3TC, the sulfur atom in the sugar moiety provided greater stability to the charge, producing fragments where the charge resided initially in the dideoxyribose (M-C₂O₂H₆).     

Applications of LC/MS and Tandem Mass Spectrometry to the Characterization of Nucleosides in Mixtures

Abstract

Liquid chromatography-mass spectrometry (LC/MS) and tandem mass spectrometry (MS/MS) provide new approaches for structural studies of nucleosides, in the nanogram range, in mixtures. Examples are given of the use of LC/MS for rapid screening of synthesis reaction mixtures, and of MS/MS for the detection and characterization of nucleoside isomers in RNA hydrolysates.