Combinatorial Chemistry & High Throughput Screening - Volume 12, Issue 2, 2009

Although rooted in physics, today mass spectrometry (MS) has become an irreplaceable tool for scientists to study the mechanisms of elemental and molecular processes occurring in nature. Due to novel MS configurations and ionisation methods, the last two decades have witnessed rapid and extensive utilisation of mass spectrometry for qualitative and quantitative purposes. Among several research fields, combinatorial chemistry has benefited from the latest MS developments by providing innovative approaches for structural characterisation of library components. However, the high sensitivity required to detect low abundance biomolecular species together with the high mass accuracy/resolving power necessary for enhancing selectivity in analysis still represent major challenges for any new development in mass spectrometry. In addition, advances in mass analysers and hardware configurations together with novel ionisation methods must also be followed by improvements in software at both the control and processing levels. Only then, more robust mass spectrometers with higher levels of automation and throughput can be delivered. Through the reviews appearing in this special issue of Combinatorial Chemistry & High Throughput Screening, we summarise the state-of-art of biological mass spectrometry. This issue brings together scientific leaders in their disciplines who approach the problems arising in mass spectrometry from different perspectives and provides a clear picture of instrumentation and applications in MS including advantages and limitations for the analysis of biomolecules. In the review by Sara Crotti and Pietro Traldi, principles of the novel atmospheric pressure ionisation techniques DESI and SACI are described including their relevance for drug analysis and metabolic profiling. Recent developments in time-of-flight (ToF) geometries for MALDI tandem mass spectrometry are reviewed by Ernst Pittenauer and Gunter Allmaier highlighting the most recent applications with various biomolecules. Simona Francese and collaborators offer a thorough summary of MALDI profiling and imaging mass spectrometry for biological tissue analysis. The impact on peptide sequencing using gas-phase fragmentation of post-translationally modified and derivatized peptides during multistage tandem mass spectrometry are addressed in the review by Jennifer Froelich and Gavin Reid. Hyphenation with liquid chromatography and the most recent developments in micro/nanoliquid chromatography interfaced with MALDI and electrospray ionisation for proteomics applications are discussed in the review by Jessica Bereszczak and Francesco Brancia. James Wright and Simon Hubbard summarise the current challenges that bioinformatics must tackle due to the high volume of complex data generated from large scale proteomics experiments. The current mass spectrometry based methods in being used to understand and characterise noncovalent protein complexes by means of electrospray ionisation are discussed by Bryan McCullogh and Simon Gaskell. Finally William Griffiths and Yuqin Wang present an overview on steroidomics in the study of neurodegenerative disease and ageing from a mass spectrometry perspective. As reported here, mass spectrometry is growing rapidly towards new directions especially for peptide/protein analysis and related research fields. This special issue provides a detailed depiction of developments/applications in mass spectrometry, which can subsequently be applied to enhance drug target identification.

The operating principles and some applications of atmospheric pressure desorption electrospray ionization (DESI) and surface activated chemical ionization (SACI) methods are described in detail. The former technique allows one to obtain information on the chemical composition (in terms of organic compounds) present on a surface of interest. The latter, SACI, provides chemical information as a result of the interaction of a vaporised solution of the analyte with a metallic surface. Both techniques typically lead to the production of abundant protonated molecules. The data available in the literature indicate that both DESI and SACI are highly promising techniques with the former giving to mass spectrometry new application fields, and the latter an increasing sensitivity and a lowering of chemical noise that, in the case of biological samples, represent a weak point in many analytical measurements.

MALDI in combination with high-energy collision-induced dissociation (CID) performed by tandem time-offlight mass spectrometry (TOF/RTOF) is a relatively new technology for the structural analysis of various classes of biomolecules as e.g., peptides, carbohydrates, glycoconjugate drugs and lipids. Fragmentation mechanisms for these classes of compounds as well as corresponding fragment ion nomenclatures based mainly on data from tandem magnetic sector mass spectrometers are summarized in this article. The major instrumental differences between the present commercially available TOF/RTOFs are compiled (e.g., ion gate, gas-collision cell, type of reflectron, etc.). Whereas peptides have been investigated by MALDI-TOF/RTOF and their CID spectra are well understood, other classes of compounds (e.g., carbohydrates or lipids) are far less well investigated. By comparing data from two different MALDI-TOF/RTOF-instruments, it becomes evident that as they are operated at rather different collision energies for CID (1 versus 20 keV) strong differences in corresponding CID spectra for the same analyte are observed, causing problems with library searches in databases as e.g., abundant peptide side-chain fragmentations mainly occurring in the 8 to 20 keV collision regime are not considered. In contrast, differences in CID spectra of carbohydrates among different TOF/RTOF instruments are less clear-cut, because the required collision energy is spread across a wide range. Especially, carbohydrate cross-ring cleavages require less collision energy in the keV-range than the corresponding peptide side-chain fragmentations. Some of these carbohydrate cross-ring fragmentations are even observed by very low energy CID (< 1 eV fragmentation amplitude). Similar observations can also be made for glycoconjugates (e.g., the drug tylosin A). The lipid class triacylglycerol needs rather high collision energies for dissociating carbon-carbon bonds based upon classical charge-remote fragmentation mechanisms. Comparison of high-energy CID-data of ESI generated triacylglycerol precursors with CID spectra from MALDI generated precursors shows different mechanisms for charge-remote fragmentations. MALDI-TOF/RTOFinstruments operated in the elevated high-energy CID mode exhibit a strong potential in structural analysis of natural and synthetic biomolecules with information often not obtainable by low energy CID.

Mass Spectrometry (MS) has a number of features namely sensitivity, high dynamic range, high resolution, and versatility which make it a very powerful analytical tool for a wide spectrum of applications spanning all the life science fields. Among all the MS techniques, MALDI Imaging mass spectrometry (MALDI MSI) is currently one of the most exciting both for its rapid technological improvements, and for its great potential in high impact bioscience fields. Here, MALDI MSI general principles are described along with technical and instrumental details as well as application examples. Imaging MS instruments and imaging mass spectrometric techniques other than MALDI, are presented along with examples of their use. As well as reporting MSI successes in several bioscience fields, an attempt is made to take stock of what has been achieved so far with this technology and to discuss the analytical and technological advances required for MSI to be applied as a routine technique in clinical diagnostics, clinical monitoring and in drug discovery.

Significant effort has been extended in recent years toward the development and application of ‘targeted’ approaches for the identification, characterization and quantitative analysis of post-translational or process-induced protein modifications, based on the multistage tandem mass spectrometry (MS/MS and MS3) fragmentation reactions of their proteolytically derived peptide ions. Although these approaches have been successfully employed to date, the development of an improved understanding of the mechanisms and other factors (e.g., proton mobility, peptide conformation, product ion structures, etc.) that influence the multistage fragmentation reactions of modified peptide ions would facilitate further advances in the field. In this review, the important role of such mechanistic studies for rationalizing the effect of posttranslational (e.g., phosphoserine- and phosphothreonine-containing peptides) and process-induced (e.g., oxidative modifications of methionine- and S-alkyl cysteine-containing peptides) protein modifications on the multistage collision induced dissociation gas-phase fragmentation reactions of proteolytically derived peptide ions are highlighted. Furthermore, recent efforts toward the development of chemical derivatization strategies for controlling and directing the gas-phase fragmentation reactions of protonated peptides toward the formation of analytically useful fragmentation pathways will be discussed, as well as the use of alternative dissociation techniques including electron capture dissociation (ECD) and electron transfer dissociation (ETD).

Hyphenation with liquid chromatography has become indispensable in mass spectrometry-based proteomics. Sample complexity together with the large variations in dynamic range can be only tackled using techniques that isolate and/or concentrate individual components prior to mass spectrometric analysis. In this review the most recent developments in micro/nanoliquid chromatography interfaced with MALDI and electrospray ionisation are discussed. Particular attention is focused on all applications related to quantitative proteomics.

Mass spectrometry has become the pre-eminent analytical method for the study of proteins and proteomes in post-genome science. The high volumes of complex spectra and data generated from such experiments represent new challenges for the field of bioinformatics. The past decade has seen an explosion of informatics tools targeted towards the processing, analysis, storage, and integration of mass spectrometry based proteomic data. In this review, some of the more recent developments in proteome informatics will be discussed. This includes new tools for predicting the properties of proteins and peptides which can be exploited in experimental proteomic design, and tools for the identification of peptides and proteins from their mass spectra. Similarly, informatics approaches are required for the move towards quantitative proteomics which are also briefly discussed. Finally, the growing number of proteomic data repositories and emerging data standards developed for the field are highlighted. These tools and technologies point the way towards the next phase of experimental proteomics and informatics challenges that the proteomics community will face.

The key strengths of electrospray over any other ionisation techniques are its soft nature and its ability to produce multiply charged ions. This combination is ideal for the study of non-covalent interactions. In this review article, we cover the basics of studying non-covalent interactions by mass spectrometry - illustrated with examples from our own and other labs - and discuss the current mass spectrometry based methods used for understanding and characterising noncovalent protein complexes.

In this mini review, the importance of experimental steroidomics in the study of neurodegenerative disease and aging is discussed. Attention is focused on just one class of lipid which is based on the cyclopentanoperhydrophenanthrene ring system. Experimental methods for steroidomic analysis are reviewed, and the potential to use these methods to diagnose disease and to gain a better understanding of neurodegenerative disorders is examined.

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