Netherlands Proteomics Center

Utrecht, Netherlands

Netherlands Proteomics Center

Utrecht, Netherlands

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Liebensteiner M.G.,Wageningen University | Pinkse M.W.H.,Technical University of Delft | Pinkse M.W.H.,Netherlands Proteomics Center | Schaap P.J.,Wageningen University | And 3 more authors.
Science | Year: 2013

Perchlorate and chlorate anions [(per)chlorate] exist in the environment from natural and anthropogenic sources, where they can serve as electron acceptors for bacteria. We performed growth experiments combined with genomic and proteomic analyses of the hyperthermophile Archaeoglobus fulgidus that show (per)chlorate reduction also extends into the archaeal domain of life. The (per)chlorate reduction pathway in A. fulgidus relies on molybdo-enzymes that have similarity with bacterial enzymes; however, chlorite is not enzymatically split into chloride and oxygen. Evidence suggests that it is eliminated by an interplay of abiotic and biotic redox reactions involving sulfur compounds. Biological (per)chlorate reduction by ancient archaea at high temperature may have prevented accumulation of perchlorate in early terrestrial environments and consequently given rise to oxidizing conditions on Earth before the rise of oxygenic photosynthesis.

Mohammed S.,University Utrecht | Mohammed S.,Netherlands Proteomics Center | Heck A.J.R.,University Utrecht | Heck A.J.R.,Netherlands Proteomics Center | Heck A.J.R.,ETH Zurich
Current Opinion in Biotechnology | Year: 2011

The multidimensional combination of strong cation exchange (SCX) chromatography and reversed phase chromatography has emerged as a powerful approach to separate peptides originating from complex samples such as digested cellular lysates or tissues before analysis by mass spectrometry, enabling the identification of over 10,000s of peptides and thousands of proteins in a single sample. Although, such multidimensional chromatography approaches are powerful, the in-depth analysis of protein post-translational modifications still requires additional sample preparation steps, involving the specific enrichment of peptides displaying the targeted modification. Here, we describe how in particular SCX chromatography can be used for the targeted analysis of important post-translational modifications, such as phosphorylation and N-terminal acetylation. Compared to other methods, SCX is less labor-intensive and more robust, and therefore likely more easily adaptable to main-stream research laboratories. © 2010 Elsevier Ltd.

Chughtai K.,FOM Institute for Atomic and Molecular Physics | Jiang L.,Johns Hopkins University | Greenwood T.R.,Johns Hopkins University | Glunde K.,Johns Hopkins University | And 2 more authors.
Journal of Lipid Research | Year: 2013

The lipid compositions of different breast tumor microenvironments are largely unknown due to limitations in lipid imaging techniques. Imaging lipid distributions would enhance our understanding of processes occurring inside growing tumors, such as cancer cell proliferation, invasion, and metastasis. Recent developments in MALDI mass spectrometry imaging (MSI) enable rapid and specific detection of lipids directly from thin tissue sections. In this study, we performed multimodal imaging of acylcarnitines, phosphatidylcholines (PC), a lysophosphatidylcholine (LPC), and a sphingomyelin (SM) from different microenvironments of breast tumor xenograft models, which carried tdTomato red fluorescent protein as a hypoxia-response element-driven reporter gene. The MSI molecular lipid images revealed spatially heterogeneous lipid distributions within tumor tissue. Four of the most-abundant lipid species, namely PC(16:0/16:0), PC(16:0/18:1), PC(18:1/18:1), and PC(18:0/18:1), were localized in viable tumor regions, whereas LPC(16:0/0:0) was detected in necrotic tumor regions. We identified a heterogeneous distribution of palmitoylcarnitine, stearoylcarnitine, PC(16:0/22:1), and SM(d18:1/16:0) sodium adduct, which colocalized primarily with hypoxic tumor regions. For the first time, we have applied a multimodal imaging approach that has combined optical imaging and MALDI-MSI with ion mobility separation to spatially localize and structurally identify acylcarnitines and a variety of lipid species present in breast tumor xenograft models. Copyright © 2013 by the American Society for Biochemistry and Molecular Biology, Inc.

Snijder J.,University Utrecht | Snijder J.,Netherlands Proteomics Center | Heck A.J.R.,University Utrecht | Heck A.J.R.,Netherlands Proteomics Center
Annual Review of Analytical Chemistry | Year: 2014

Analysis of the size and mass of nanoparticles, whether they are natural biomacromolecular or synthetic supramolecular assemblies, is an important step in the characterization of such molecular species. In recent years, electrospray ionization (ESI) has emerged as a technology through which particles with masses up to 100 MDa can be ionized and transferred into the gas phase, preparing them for accurate mass analysis. Here we review currently used methodologies, with a clear focus on native mass spectrometry (MS). Additional complementary methodologies are also covered, including ion-mobility analysis, nanomechanical mass sensors, and charge-detection MS. The literature discussed clearly demonstrates the great potential of ESI-based methodologies for the size and mass analysis of nanoparticles, including very large naturally occurring protein assemblies. The analytical approaches discussed are powerful tools in not only structural biology, but also nanotechnology. © Copyright ©2014 by Annual Reviews. All rights reserved.

Di Palma S.,University Utrecht | Di Palma S.,Netherlands Proteomics Center | Mohammed S.,University Utrecht | Mohammed S.,Netherlands Proteomics Center | And 2 more authors.
Nature Protocols | Year: 2012

Multidimensional liquid chromatography (LC) combined with mass spectrometry (MS) has become a standard technique in proteomics to reduce sample complexity and to tackle the dynamic range in protein abundance. Fractionation is necessary to obtain a comprehensive analysis of complex biological samples such as tissue and mammalian cell lines. However, extensive fractionation comes at the expense of sample loss, presenting a bottleneck in the analysis of limited amounts of material. In this protocol, we describe a two-dimensional chromatographic strategy based on a combination of hydrophilic interaction liquid chromatography (HILIC; with a zwitterionic packing material, ZIC-cHILIC) and reversed-phase chromatography, which allows proteomic analyses with minimal sample loss. Experimental aspects related to obtaining maximum recovery are discussed, including how to optimally prepare samples for this system. Examples involving protein lysates originating from cultured cell lines and cells sorted by flow cytometry are used to show the power, sensitivity and versatility of the technique. Once the ZIC-cHILIC fractionation system has been optimized and standardized, this protocol requires ∼5-6 d, including sample preparation and fraction analysis. © 2012 Nature America, Inc. All rights reserved.

Uetrecht C.,University Utrecht | Uetrecht C.,Netherlands Proteomics Center | Uetrecht C.,Uppsala University | Heck A.J.R.,University Utrecht | Heck A.J.R.,Netherlands Proteomics Center
Angewandte Chemie - International Edition | Year: 2011

Over a century since its development, the analytical technique of mass spectrometry is blooming more than ever, and applied in nearly all aspects of the natural and life sciences. In the last two decades mass spectrometry has also become amenable to the analysis of proteins and even intact protein complexes, and thus begun to make a significant impact in the field of structural biology. In this Review, we describe the emerging role of mass spectrometry, with its different technical facets, in structural biology, focusing especially on structural virology. We describe how mass spectrometry has evolved into a tool that can provide unique structural and functional information about viral-protein and protein-complex structure, conformation, assembly, and topology, extending to the direct analysis of intact virus capsids of several million Dalton in mass. Mass spectrometry is now used to address important questions in virology ranging from how viruses assemble to how they interact with their host. © 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Altelaar A.F.M.,University Utrecht | Altelaar A.F.M.,Netherlands Proteomics Center | Munoz J.,University Utrecht | Munoz J.,Netherlands Proteomics Center | And 3 more authors.
Nature Reviews Genetics | Year: 2013

Next-generation sequencing allows the analysis of genomes, including those representing disease states. However, the causes of most disorders are multifactorial, and systems-level approaches, including the analysis of proteomes, are required for a more comprehensive understanding. The proteome is extremely multifaceted owing to splicing and protein modifications, and this is further amplified by the interconnectivity of proteins into complexes and signalling networks that are highly divergent in time and space. Proteome analysis heavily relies on mass spectrometry (MS). MS-based proteomics is starting to mature and to deliver through a combination of developments in instrumentation, sample preparation and computational analysis. Here we describe this emerging next generation of proteomics and highlight recent applications. © 2013 Macmillan Publishers Limited.

Rosati S.,University Utrecht | Rosati S.,Netherlands Proteomics Center | Yang Y.,University Utrecht | Yang Y.,Netherlands Proteomics Center | And 4 more authors.
Nature Protocols | Year: 2014

The molecular complexity of biopharmaceuticals puts severe demands on the bioanalytical techniques required for their comprehensive structural characterization. Mass spectrometry (MS) has gained importance in the analysis of biopharmaceuticals, taking different complementary approaches ranging from peptide-based sequencing to direct analysis of intact proteins and protein assemblies. In this protocol, we describe procedures optimized to perform the analysis of monoclonal antibodies (mAbs) at the intact protein level under pseudo-native conditions, using native MS. Some of the strengths of native MS in the analysis of biopharmaceuticals are its analysis speed, sensitivity and specificity: for most experiments, the whole protocol requires one working day, whereby tens of samples can be analyzed in a multiplexed manner, making it suitable for high-throughput analysis. This method can be used for different applications such as the analysis of mixtures of mAbs, drug-antibody conjugates and the analysis of mAb post-translational modifications, including the qualitative and quantitative analysis of mAb glycosylation. © 2014 Nature America, Inc. All rights reserved.

Altelaar A.F.M.,University Utrecht | Altelaar A.F.M.,Netherlands Proteomics Center | Heck A.J.R.,University Utrecht | Heck A.J.R.,Netherlands Proteomics Center
Current Opinion in Chemical Biology | Year: 2012

Here we review recent developments and trends in sample preparation, pre-fractionation, chromatography and mass spectrometry contributing towards the ultra-sensitive global analysis of proteins. Highly sensitive MS-based proteomics is not only beneficiary for the proteome analysis of single cells, an aim which is getting into reach, but also clearly relevant for the analysis of (a) subcellular organelles, (b) specific low-abundant cell-types such as adult stem cells, and (c) smaller but more homogeneous cell populations sorted or dissected from (diseased) tissue. © 2011 Elsevier Ltd.

Rose R.J.,University Utrecht | Rose R.J.,Netherlands Proteomics Center | Damoc E.,Thermo Fisher Scientific | Denisov E.,Thermo Fisher Scientific | And 3 more authors.
Nature Methods | Year: 2012

The analysis of intact protein assemblies in native-like states by mass spectrometry offers a wealth of information on their biochemical and biophysical properties. Here we show that the Orbitrap mass analyzer can be used to measure protein assemblies of molecular weights approaching one megadalton with sensitivity down to the detection of single ions. Minor instrumental modifications enabled the measurement of various protein assemblies with outstanding mass-spectral resolution. © 2012 Nature America, Inc. All rights reserved.

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