Since the introduction of travelling wave (T-Wave)-based ion mobility in 2007 a large number of research laboratories have embraced the technique, particularly those working in the field of structural biology. The development of software to process the data generated from this technique, however, has been limited. We present a novel software package that enables the processing of T-Wave ion mobility data. The program can deconvolute components in a mass spectrum and uses this information to extract corresponding arrival time distributions (ATDs) with minimal user intervention. It can also be used to automatically create a collision cross section (CCS) calibration and apply this to subsequent files of interest. A number of applications of the software, and how it enhances the information content extracted from the raw data, are illustrated using model proteins.

Members of the RASSF family (RASSF1-10) have been identified as candidate tumour suppressors that are frequently downregulated by promoter hypermethylation in cancers. These proteins carry a common Ras-association (RA) and SARAH domain (RASSF1-6) that can potentially bind Ras oncoproteins and mediate protein–protein interactions with other SARAH domain proteins. However, there is a notable lack of comparative characterisation of the RASSF family, as well as molecular and structural information that facilitate their tumour suppressive functions. As part of our comparative analysis, we modelled the RA and SARAH domains of the RASSF members based on existing structures and predicted their potential interactions. These in silico predictions were compared to in vitro interaction studies with Ras and MST kinase (a SARAH domain-containing protein). Our data shows a diversity of interaction within the RASSF family RA domain, whereas the SARAH domain-mediated interactions for RASSF1-6 are consistent with the predictions. This suggests that different members, despite shared general architecture, could have distinct functional properties. Additionally, we identify a new interacting partner for MST kinase in the form of RASSF7. Current data supports an interaction model where RASSF serves as an adaptor for the assembly of multiple protein complexes and further functional interactions, involving MST kinases and other SARAH domain proteins, which could be regulated by Ras.

The malaria parasite exports proteins across its plasma membrane and a surrounding parasitophorous vacuole membrane, into its host erythrocyte. Most exported proteins contain a Host Targeting motif (HT motif) that targets them for export. In the parasite secretory pathway, the HT motif is cleaved by the protease plasmepsin V, but the role of the newly generated N-terminal sequence in protein export is unclear. Using a model protein that is cleaved by an exogenous viral protease, we show that the new N-terminal sequence, normally generated by plasmepsin V cleavage, is sufficient to target a protein for export, and that cleavage by plasmepsin V is not coupled directly to the transfer of a protein to the next component in the export pathway. Mutation of the fourth and fifth positions of the HT motif, as well as amino acids further downstream, block or affect the efficiency of protein export indicating that this region is necessary for efficient export. We also show that the fifth position of the HT motif is important for plasmepsin V cleavage. Our results indicate that plasmepsin V cleavage is required to generate a new N-terminal sequence that is necessary and sufficient to mediate protein export by the malaria parasite.

This paper presents the structure of MsAcg (MSMEG_5246), a Mycobacterium smegmatis homologue of Mycobacterium tuberculosis Acg (Rv2032) in its reduced form at 1.6 Å resolution using x-ray crystallography. Rv2032 is one of the most induced genes under the hypoxic model of tuberculosis dormancy. The Acg family turns out to be unusual flavin mononucleotide (FMN)-binding proteins that have probably arisen by gene duplication and fusion from a classical homodimeric nitroreductase such that the monomeric protein resembles a classical nitroreductase dimer but with one active site deleted and the other active site covered by a unique lid. The FMN cofactor is not reduced by either NADH or NADPH, but the chemically reduced enzyme is capable of reduction of nitro substrates, albeit at no kinetic advantage over free FMN. The reduced enzyme is rapidly oxidized by oxygen but without any evidence for a radical state commonly seen in oxygen-sensitive nitroreductases. The presence of the unique lid domain, the lack of reduction by NAD(P)H, and the slow rate of reaction of the chemically reduced protein raises a possible alternative function of Acg proteins in FMN storage or sequestration from other biochemical pathways as part of the bacteria's adaptation to a dormancy state.

The proteome of the bacterium Methylocella silvestris has been characterized using reversed phase ultra high pressure liquid chromatography (UPLC) and two-dimensional reversed phase (high pH)–reversed phase (low pH) UPLC prior to mass spectrometric analysis. Variations in protein expression levels were identified with the aid of label-free quantification in a study of soluble protein extracts from the organism grown using methane, succinate, or propane as a substrate. The number of first dimensional fractionation steps has been varied for 2D analyses, and the impact on data throughput and quality has been demonstrated. Comparisons have been made regarding required experimental considerations including total loading of biological samples required, instrument time, and resulting data file sizes. The data obtained have been evaluated with respect to number of protein identifications, confidence of assignments, sequence coverage, relative levels of proteins, and dynamic range. Good qualitative and quantitative agreement was observed between the different approaches, and the potential benefits and limitations of the reversed phase–reversed phase UPLC technique in label-free analysis are discussed. A preliminary screen of the protein regulation data has also been performed, providing evidence for a possible propane assimilation route.

The functional design and application of a data-independent LC-MS precursor and product ion repository for protein identification, quantification, and validation is conceptually described. The ion repository was constructed from the sequence search results of a broad range of discovery experiments investigating various tissue types of two closely related mammalian species. The relative high degree of similarity in protein complement, ion detection, and peptide and protein identification allows for the analysis of normalized precursor and product ion intensity values, as well as standardized retention times, creating a multidimensional/orthogonal queryable, qualitative, and quantitative space. Peptide ion map selection for identification and quantification is primarily based on replication and limited variation. The information is stored in a relational database and is used to create peptide- and protein-specific fragment ion maps that can be queried in a targeted fashion against the raw or time aligned ion detections. These queries can be conducted either individually or as groups, where the latter affords pathway and molecular machinery analysis of the protein complement. The presented results also suggest that peptide ionization and fragmentation efficiencies are highly conserved between experiments and practically independent of the analyzed biological sample when using similar instrumentation. Moreover, the data illustrate only minor variation in ionization efficiency with amino acid sequence substitutions occurring between species. Finally, the data and the presented results illustrate how LC-MS performance metrics can be extracted and utilized to ensure optimal performance of the employed analytical workflows.

Type IV secretion (T4S) systems mediate the transfer of proteins and DNA across the cell envelope of bacteria. These systems play important roles in bacterial pathogenesis and in horizontal transfer of antibiotic resistance. The VirB4 ATPase of the T4S system is essential for both the assembly of the system and substrate transfer. In this article, we present the crystal structure of the C-terminal domain of Thermoanaerobacter pseudethanolicus VirB4. This structure is strikingly similar to that of another T4S ATPase, VirD4, a protein that shares only 12% sequence identity with VirB4. The VirB4 domain purifies as a monomer, but the full-length protein is observed in a monomer-dimer equilibrium, even in the presence of nucleotides and DNAs. We also report the negative stain electron microscopy structure of the core complex of the T4S system of the Escherichia coli pKM101 plasmid, with VirB4 bound. In this structure, VirB4 is also monomeric and bound through its N-terminal domain to the core’s VirB9 protein. Remarkably, VirB4 is observed bound to the side of the complex where it is ideally placed to play its known regulatory role in substrate transfer.

In conformational diseases, native protein conformers convert to pathological intermediates that polymerize. Structural characterization of these key intermediates is challenging. They are unstable and minimally populated in dynamic equilibria that may be perturbed by many analytical techniques. We have characterized a forme fruste deficiency variant of α1-antitrypsin (Lys154Asn) that forms polymers recapitulating the conformer-specific neo-epitope observed in polymers that form in vivo. Lys154Asn α1-antitrypsin populates an intermediate ensemble along the polymerization pathway at physiological temperatures. Nuclear magnetic resonance spectroscopy was used to report the structural and dynamic changes associated with this. Our data highlight an interaction network likely to regulate conformational change and do not support the recent contention that the disease-relevant intermediate is substantially unfolded. Conformational disease intermediates may best be defined using powerful but minimally perturbing techniques, mild disease mutants, and physiological conditions.

The HtrA protein family combines chaperone and protease activities and is essential for protein quality control in many organisms. Whereas the mechanisms underlying the proteolytic function of HtrA proteins are well characterized, their chaperone activity remains poorly understood. Here we describe cryo-EM structures of Escherichia coli DegQ in its 12- and 24-mer states in complex with model substrates, providing a structural model of HtrA chaperone action. Up to six lysozyme substrates bind inside the DegQ 12-mer cage and are visualized in a close-to-native state. An asymmetric reconstruction reveals the binding of a well-ordered lysozyme to four DegQ protomers. DegQ PDZ domains are located adjacent to substrate density and their presence is required for chaperone activity. The substrate-interacting regions appear conserved in 12- and 24-mer cages, suggesting a common mechanism of chaperone function.

The analysis of glycosylation from native biological sources is often frustrated by the low abundances of available material. Here, ion mobility combined with electrospray ionization mass spectrometry have been used to extract the spectra of N-glycans released with PNGase F from a serial titration of recombinantly expressed envelope glycoprotein, gp120, from the human immunodeficiency virus (HIV). Analysis was also performed on gp120 expressed in the α-mannosidase inhibitor, and in a matched mammalian cell line deficient in GlcNAc transferase I. Without ion mobility separation, ESI spectra frequently contained no observable ions from the glycans whereas ions from other compounds such as detergents and residual buffer salts were abundant. After ion mobility separation on a Waters T-wave ion mobility mass spectrometer, the N-glycans fell into a unique region of the ion mobility/m/z plot allowing their profiles to be extracted with good signal:noise ratios. This method allowed N-glycan profiles to be extracted from crude incubation mixtures with no clean-up even in the presence of surfactants such as NP40. Furthermore, this technique allowed clear profiles to be obtained from sub-microgram amounts of glycoprotein. Glycan profiles were similar to those generated by MALDI-TOF MS although they were more susceptible to double charging and fragmentation. Structural analysis could be accomplished by MS/MS experiments in either positive or negative ion mode but negative ion mode gave the most informative spectra and provided a reliable approach to the analysis of glycans from small amounts of glycoprotein.

A combination of ion mobility and mass spectrometry methods was used to characterize the molecular shape of the protein calmodulin (CaM) and its complexes with calcium and a number of peptide ligands. CaM, a calcium-binding protein composed of 148 amino acid residues, was found by X-ray crystallography to occur both in a globular shape and in the shape of an extended dumbbell. Here, it was found, as solutions of Cam and Cam complexes were sprayed into the solvent-free environment of the mass spectrometer, that major structural feature,., of the molecule and the stoichiometry of the units constituting a complex in Solution were preserved in the desolvation process. Two types of Cam structures were observed in Our experiments: a compact and all extended form of Cam with measured cross sections in near-perfect agreement with those calculated for the known globular and extended dumbbell X-ray geometries. Calcium-free Solutions yielded predominantly all extended Cam conformation. Ca-n(2+)-CaM complexes were observed in Calcium-containing solutions, n = 0-4, with the population of the compact conformation increasing relative to the elongated conformation as n increases. For n = 4, a predominantly compact globular conformation was observed. Solutions containing the peptide CaMKII290-309, the CaM target domain of the Ca2+/Calmodulin-dependent protein kinase II (CaMKII) enzyme, yielded predominantly globular Ca-4(2+)-CaM-CaMKII290-309 complexes. Similar results were obtained with the 26-residue peptide melittin. For the 14-residue C-terminal melittin fragment, oil the other hand, formation of both a 1: 1 and a 1:2 CaM-peptide complex was detected. Oil the basis of the entirety of our results, we Conclude that the collapse of extended (dumbbell-like) CaM structures into more compact globular Structures Occurs upon specific binding Of four calcium ions. Furthermore, this calcium-induced structural Collapse of Cam appears to be a prerequisite for formation of a particularly stable CaM-peptide complex involving peptides long enough to be engaged in interactions with both lobes of CaM.

Poly(2-hydroxyethyl methacrylate) (poly(HEMA)) has been characterised by means of electrospray ionisation-mass spectrometry/mass spectrometry (ESI-MS/MS), in order to evaluate this technique for the generation of end group information. Low energy collision-induced dissociation (CID) data from poly(HEMA) enabled information on both end groups of the polymer chain to be gleaned, in a similar fashion to that proposed previously for other methacrylate polymer systems. Exact-mass CID information was employed to aid the understanding of the dissociation mechanism of the polymer Some additional fragmentation pathways, compared to other methacrylate polymer systems, are proposed. An example of how software can aid the interpretation of the MS/MS data is also shown (C) 2010 Elsevier Ltd All rights reserved.

The prion protein (PrP) is implicitly involved in the pathogenesis of transmissible spongiform encephalopathies (TSEs). The conversion of normal cellular PrP (PrPC), a protein that is predominantly alpha-helical, to a beta-sheet-rich isoform (PrPSc), which has a propensity to aggregate, is the key molecular event in prion diseases. During its short life span, PrP can experience two different: pH environments; a mildly acidic environment, whilst cycling within the cell, and a neutral pH when it is glycosyl phosphatidylinositol (GPI)-anchored to the cell membrane. Ion mobility (IM) combined with mass spectrometry has been employed to differentiate between two conformational isoforms of recombinant Syrian hamster prion protein (SHaPrP). The recombinant proteins studied were a-helical SHaPrP(90-231) and beta-sheet-rich SHaPrP(90-231) at pH 5.5 and pH 7.0. The recombinant proteins have the same nominal mass-to-charge ratio (m/z) but differ in their secondary and tertiary structures. A comparison of traveling-wave (T-Wave) ion mobility and drift cell ion mobility (DCIM) mass spectrometry estimated and absolute cross-sections showed an excellent agreement between the two techniques. The use of T-Wave ion mobility as a shape-selective separation technique enabled differentiation between the estimated cross-sections and arrival time distributions (ATDs) of alpha-helical SHaPrP(90-231) and beta-sheet-rich SHaPrP(90-231) at pH 5.5. No differences in cross-section or ATD profiles were observed between the protein isoforms at pH 7.0. The findings have potential implications for a new ante-mortem screening assay, in bodily fluids, for prion misfolding diseases such as TSEs. (J Am Soc Mass Spectrom 2010, 21, 845-854) (C) 2010 American Society for Mass Spectrometry

Phosphorylation is one the most studied and important post translational modifications. Nano electrospray mass spectrometry coupled with traveling wave (T-Wave)-based ion mobility has been used to filter for phosphorylated peptides in tryptic protein digests. T-Wave parameters have been optimized to maximize the separation between phosphorylated and non-phosphorylated peptides. A method to calibrate the T-Wave device, to provide estimates of collision cross sections, is presented, and these estimates are in excellent agreement with values obtained on drift cell instrumentation. Phosphorylated peptides have smaller cross sections which enables their separation from non-phosphorylated peptides of the same m/z. Post-mobility fragmentation is used to obtain the primary sequence for peptides of interest. This approach is shown to have potential as an additional screen for phosphorylated peptides, where up to 40% of observed peptides can be eliminated from the study.

Hemoglobin (Hb) is a tetrameric noncovalent complex consisting of two alpha- and two beta-globin chains each associated with a heme group. Its exact assembly pathway is a matter of debate. Disorders of hemoglobin are the most common inherited disorders and subsequently the molecule has been extensively studied. This work attempts to further elucidate the structural properties of the hemoglobin tetramer and its components. Gas-phase conformations of hemoglobin tetramers and their constituents were investigated by means of traveling-wave ion mobility mass spectrometry. Sickle (HbS) and normal (HbA) hemoglobin molecules were analyzed to determine whether conformational differences in their quaternary structure could be observed. Rotationally averaged collision cross sections were estimated for tetramer, dimer, apo-, and holo-monomers with reference to a protein standard with known cross sections. Estimates of cross section obtained for the tetramers were compared to values calculated from X-ray crystallographic structures. HbS was consistently estimated to have a larger cross section than that of HbA, comparable with values obtained from X-ray crystallographic structures. Nontetrameric species observed included apo- and holo- forms of alpha- and beta-monomers and heterodimers: alpha- and beta-monomers in both apo- and holo- forms were found to have similar cross sections, suggesting they maintain a similar fold in the gas phase in both the presence and the absence of heme. Heme-deficient dimer, observed in the spectrum when analyzing commercially prepared Hb, was not observed when analyzing fresh blood. This implies that holo-alpha-apo-beta is not an essential intermediate within the Hb assembly pathway, as previously proposed. (J Am Soc Mass Spectrom 2009, 20, 625-631) (C) 2009 Published by Elsevier Inc. on behalf of American Society for Mass Spectrometry

The proteome of the recently discovered bacterium Methyiocella silvestris has been characterized using three profiling and comparative proteomics approaches. The organism has been grown on two different substrates enabling variations in protein expression to be identified. The results obtained using the experimental approaches have been compared with respect to number of proteins identified, confidence in identification, sequence coverage and agreement of regulated proteins. The sample preparation, instrumental time and sample loading requirements of the differing experiments are compared and discussed. A preliminary screen of the protein regulation results for biological significance has also been performed.

The methodology developed in the research presented herein makes use of chaotropic solvents to gently dissociate subunits from an intact macromolecular complex and subsequently allows for the measurement of collision cross section (CCS) for both the recombinant (R-eIF3k) and solvent dissociated form of the subunit (S-eIF3k). In this particular case, the k Subunit from the eukaryotic initiation factor 3 (eIF3) was investigated in detail. Experimental and theoretical CCS values show both the recombinant and solvent disrupted forms of the protein to be essentially the same. The ultimate goal of the project is to structurally characterize all the binding partners of eIF3, determine which subunits interact directly, and investigate how subunits may change conformation when they form complexes with other proteins. Research presented herein is the first report showing retention of solution conformation of a protein as evidenced by CCS measurements of both recombinant and solvent disrupted versions of the same protein. (J Am Soc Mass Spectrom 2009, 20, 1699-1706) (C) 2009 Published by Elsevier Inc. on behalf of American Society for Mass Spectrometry

The three-dimensional conformation of a protein is central to its biological function. The characterisation of aspects of three-dimensional protein structure by mass spectrometry is an area of much interest as the gas-phase conformation, in many instances, can be related to that of the solution phase. Travelling wave ion mobility mass spectrometry (TWIMS) was used to investigate the biological significance of gas-phase protein structure. Protein standards were analysed by TWIMS under denaturing and near-physiological solvent conditions and cross-sections estimated for the charge states observed. Estimates of collision cross-sections were obtained with reference to known standards with published cross-sections. Estimated cross-sections were compared with values from published X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy structures. The cross-section measured by ion mobility mass spectrometry varies with charge state, allowing the unfolding transition of proteins in the gas phase to be studied. Cross-sections estimated experimentally for proteins studied, for charge states most indicative of native structure, are in good agreement with measurements calculated from published X-ray and NMR structures. The relative stability of gas-phase structures has been investigated, for the proteins studied, based on their change in cross-section with increase in charge. These results illustrate that the TWIMS approach can provide data on three-dimensional protein structures of biological relevance. Copyright (C) 2008 John Wiley & Sons, Ltd.

The end-group functionalisation of a series of poly(propylene glycol)s has been characterised by means of electrospray ionisation-tandem mass spectrometry (ESI-MS/MS). A series of peaks with mass-to-charge ratios that are close to that of the precursor ion were used to generate information on the end-group functionalities of the poly(propylene glycol)s. Fragment ions resulting from losses of both of the end groups were noted from some of the samples. An example is presented of how software can be used to significantly reduce the length of time involved in data interpretation (which is typically the most time-consuming part of the analysis).

Ion mobility (IM) combined with tandem mass spectrometry (MS/MS) has been employed to separate and differentiate between polyether oligomers with the same nominal molecular weights. Poly(ethylene glycol)s with the same nominal mass-to-charge ratio (m/z), but with differing structures, were separated using ion mobility. IMMS/MS data were able to aid identification of the backbone and end groups of the four individual polyethers in the two sets of isobaric mixtures. The MS/MS data from the resolved oligomers enabled a detailed structural characterization of the polyether mixtures to be completed in one experiment.

In this study, an MS-based proteomics approach to characterizing the virion structural proteins of the novel marine 'photosynthetic' phage S-PM2 is presented. The virus infects ecologically important cyanobacteria of the genus Synechococcus that make a substantial contribution to primary production in the oceans. The S-PM2 genome encodes 236 ORFs, some of which exhibit similarity to known phage virion structural proteins, but the majority (54%) show no detectable homology to known proteins from other organisms. Using public and in-house bioinformatics; tools the proteome of S-PM2 was predicted and a database compatible with MS-based search engines was constructed. S-PM2 virion proteins were resolved by SIDS-PAGE, excised, tryptically digested and analysed by LC-ESI-MS/MS. The resulting MS data were searched against the database. A parallel control study was undertaken on the well-characterized coliphage T4 in order to assess the sensitivity and efficiency of this approach. In total, 11 of the 15 S-PM2 proteins, predicted to be virion proteins by bioinformatics approaches, were confirmed as such, together with the identification of a further 12 novel structural proteins. In the case of T4, 24 of the 39 known virion structural proteins were identified, including the major tail-fibre proteins. This approach has wide-ranging applicability and can be applied to any novel organism whose genome encodes ORFs with few detectable homologies in the public databases.

A rapid method for the characterisation of polyglycol esters and ethers is described which uses accurate mass desorption electrospray ionisation (DESI) quadrupole time-of-flight mass spectrometry (Q-ToFMS). The results are combined with newly developed software which aids the interpretation of product ions produced using collision-induced dissociation (CID) of selected precursor ions. The poly(ethylene glycol) (PEG) samples analysed were PEG dibenzoate, PEG monooleate, PEG butyl ether, PEG bis(2-ethyl hexanoate) and PEG diacrylate. Lithium metal was used for cationisation of the PEG oligomers since it yielded the most useful structural information compared with other group I metals. The full scan mass spectra and product ion mass spectra were all obtained in <5 s. Interpretation of the MS/MS product ion spectra, using the product ion interpretation software which incorporates previously developed fragmentation rules, was carried out in <1 s. Copyright (C) 2007 John Wiley & Sons, Ltd.

Novel software has been developed to aid the interpretation of tandem mass spectrometry (MS/MS) data from synthetic polymers. The software is particularly focused toward aiding the end-group determination of these materials by significantly speeding up the interpretation process. This allows information on the initiator and/or chain transfer agents, used to generate the polymer, and the mechanism of termination to be inferred from the data much more rapidly. The software allows the validity of hypothesized structures to be rapidly tested by automatically annotating the data file using previously proposed fragmentation rules for synthetic polymers. Low-energy collision-induced dissociation (CID) data from methacrylate, styrene, and polyether oligomers are used as example data for the software. Exact-mass CID information was used to aid the understanding of the dissociation mechanism of the polymers. The software can use exact-mass data to provide more confidence in the results. The MS/MS results indicate that the fragmentation pathways are those previously proposed for these polymers.

Ion mobility coupled with mass spectrometry has evolved into a powerful analytical technique for investigating the gas-phase structures of biomolecules. Here we present the mobility separation of some peptide and protein ions using a new hybrid quadrupole/travelling wave ion mobility separator/orthogonal acceleration time-of-flight instrument. Comparison of the mobility data obtained from the relatively new travelling wave separation device with data obtained using various other mobility separators demonstrate that whilst the mobility characteristics are similar, the new hybrid instrument geometry provides mobility separation without compromising the base sensitivity of the mass spectrometer. This capability facilitates mobility studies of samples at analytically significant levels. (C) 2006 Elsevier B.V. All rights reserved.

Ion mobility has emerged as an important technique for determining biopolymer conformations in solvent free environments. These experiments have been nearly exclusively performed on home built systems. In this paper we describe modifications to a commercial high performance mass spectrometer, the Waters UK "Uhima" Q-Tof, that allows high sensitivity measurement of peptide and protein cross sections. Arrival time distributions are obtained for a series of peptides (bradykinin, LHRH, substance P, bombesin) and proteins (bovine and equine cytochrome c, myoglobin, alpha-lactalbumin) with good agreement found with literature cross sections where available. In complex ATD's, mass spectra can be obtained for each feature confirming assignments. The increased sensitivity of the commercial instrument is retained along with the convenience of the data system, crucial features for analysis of protein misfolding systems. (C) 2004 Elsevier B.V. All rights reserved.

Native, metastable serpins inherently tend to undergo stabilizing conformational transitions in mechanisms of health (e.g., enzyme inhibition) and disease (serpinopathies). This intrinsic tendency is modifiable by ligand binding, thus structure-based drug design is an attractive strategy in the serpinopathies. This can be viewed as a labor-intensive approach, and historically, its intellectual attractiveness has been tempered by relatively limited success in development of drugs reaching clinical practice. However, the increasing availability of a range of powerful experimental systems and higher-throughput techniques is causing academic and early-stage industrial pharmaceutical approaches to converge. In this review, we outline the different systems and techniques that are bridging the gap between what have traditionally been considered distinct disciplines. The individual methods are not serpin-specific. Indeed, many have only recently been applied to serpins, and thus investigators in other fields may have greater experience of their use to date. However, by presenting examples from our work and that of other investigators in the serpin field, we highlight how techniques with potential for automation and scaling can be combined to address a range of context-specific challenges in targeting the serpinopathies.

Widely used in medical research, pharmaceutical and fine chemicals industries, biological and physical sciences, and security and environmental agencies, mass spectrometry techniques are continually under development. In Practical Aspects of Trapped Ion Mass Spectrometry: Volume V, Applications of Ion Trapping Devices, an international panel of authors presents a world-wide view of the practical aspects of recent progress using trapped ion devices. In contrast to previous texts, which have concentrated generally on a single or limited range of ion trapping techniques, a key feature of this compilation of contributions is its coverage of all the ion trapping techniques currently in use. Spanning sixteen chapters, the text examines:

• Ion/neutral and ion/ion reactions
• Structural characterization of proteins and peptides using quadrupole ion trap mass spectrometry, Fourier transform - ion cyclotron resonance (FT-ICR) mass spectrometry, and traveling wave ion mobility mass spectrometry
• Ion spectroscopy and electron diffraction
• Conformational analysis of protein isobaric charge states
• Practical examples of trapped-ion technology that reflect the wide diversity of applications of trapped-ion devices

This text is the first detailed account of the application of new and established mass spectrometric techniques utilizing trapped or confined ions for prolonged investigation and increased sensitivity. Each chapter contains complete references and utilizes a consistent format and writing style, with all terms, acronyms, procedures, and equations thoroughly explained. The strong editorial input to the diverse sections enables readers to readily appreciate the commonalities of topics ranging from theory of instrument operation to proteins, flavonoids, atomic clocks, and single ion mass spectrometry.