Senior Researchers

Alina Astefanei
Research

SAWN-MS, LC-SWATH-MS, cultural heritage, forensic, proteomics

Alina obtained her PhD degree in Analytical Chemistry from the University of Barcelona. Her research concerned the exciting field of fullerene nanoparticles. She has developed analytical methodologies based on AF4-MALS, CE and LC-MS that enabled their analysis and characterisation in environmental and cosmetic samples.
Currently, Alina works in the Biomolecular Systems Analytics at HIMS-UvA. Her research involves the employment of LC-SWATH-MS strategies to increase the speed of analysis of protein digest and also the development of novel ionization techniques for mass spectrometry for various fields. Alina pioneered several applications using SAWN-MS to study artworks, forensic explosives and drugs of abuse in environmental waters. For this, she collaborates with other institutes within UvA, the RCE, NFI, and several industrial partners.

Contact details

Alina Astefanei, PhD
University of Amsterdam
Van 't Hoff Institute for Molecular Sciences (HIMS)

Postal address
Postbus 94157
1090 GD Amsterdam

Visiting address
Science Park 904
1098 XH Amsterdam

email: A.Astefanei@uva.nl
researchgate.net/profile/Alina_Astefanei

Luitzen de Jong
Research

Mass spectrometric analysis of in vivo cross-linked protein complexes
After my retirement in November 2012, I continued as a guest worker my research activities arising from the work described in the thesis by Hansuk Buncherd entitled “Development of enrichment methods for cross-linked peptides to study the dynamic topology of large protein complexes by mass spectrometry”, defended December 2013. The aim of the project is to obtain a work flow enabling application of the technology developed in this thesis to an in vivo cross-linking approach of exponentially growing bacteria directly in the growth medium. This project in the field of analytical (bio)chemistry has a multidisciplinary nature and is carried out in close collaboration with microbiologists, organic chemists and mass spectrometrists.

For cross-linking we developed bis(succinimidy)-3-azidomethyl-glurtarate (BAMG). This cross-linker reacts with ε-aminogroups of juxtaposed lysine residues on the surface of proteins and protein complexes. Upon digestion following the cross-linking reaction, cross-linked peptides are generated along with an excess of unmodified peptides.

dejong-researchAim of the mass spectrometric analysis is to determine which lysine residues from which proteins became covalently linked during reaction with BAMG. This information can reveal the topology (i.e., spatial arrangement of composing proteins) of biological assemblies and is a prerequisite to understand the molecular mechanisms of complex processes, like transcription, translation and cell division. These are examples of biological processes accomplished by large protein complexes. The azido group in the spacer of BAMG enables isolation of cross-linked peptides and tremendously facilitates their identification by mass spectrometry and database searching.

The choice to use in vivo cross-linking directly in the growth medium is based on the notion that in vivo protein-protein interactions can be easily interrupted upon protein isolation or upon harvesting bacteria from the medium by centrifugation or filtering.

For growth, bacteria require, a.o., certain amines in the growth medium as a source of nitrogen. For in vivo cross-linking of exponentially growing bacteria, a medium is needed with a low concentration of compounds with amine groups, to prevent reaction with the cross-linker, but still enables fast growth. At present we are optimizing conditions for (i) growth, (ii) cross-linking directly in a bacterium culture, (iii) protein extraction and (iv), enrichment of certain cross-linked protein complexes and their digestion. The focus will be on protein complexes involved in cell division.

The work will shed new light on the molecular mechanisms underlying the cleavage process whereby one mother cell gives rise to two daughter cells. This knowledge can give clues towards the development of new antibiotics. The work flow to be developed can also be used to study several other processes and organizing principles in living cells.

A key reference is : Buncherd H, Roseboom W, De Koning LJ, De Koster CG, De Jong L (2014). A gas phase cleavage reaction of cross-linked peptides for protein complex topology studies by peptide fragment fingerprinting from large sequence database. J Proteomics 108, 65-77.

Contact details

de-jong-luitzen

Luitzen de Jong, PhD
University of Amsterdam
Swammerdam Institute of Life Sciences (SILS)
Mass Spectrometry of Biomacromolecules

Visiting address
Sciencepark 904 (room C2.267)
1098 XH Amsterdam

email: l.dejong@uva.nl

Eva de Rijke
Research

I work as a labmanager in the soil and environmental chemistry laboratories of the Institute of Biodiversity and Ecosystem Dynamics (IBED).
Available equipment in the lab to determine the (in)organic composition of soils and water includes ICP/OES for (trace) metals, an auto analyzer for the detection of anions such as nitrates and phosphates, CHN and TOC analyzers to quantify organic substances (C, N) in solid and liquid samples. In relation to this, we have facilities for soil texture analysis, soil physical properties, as well as several selective extraction methods.
To determine the composition of natural and anthropogenic organic substances we have a well equipped chromatography lab that includes GC/MS, pyrolysis-GC/MS, LC/MS and LC/MS/MS.
Furthermore we have two isotope ratio mass spectrometers (IRMS) for the analysis of stable isotopes (13C, 15N, 2H, 18O) in solid, liquid, gaseous samples, which are coupled to TOC, (high temperature) combustion and pyrolysis (solids/liquids) elemental analyzers, or GC to allow for compound specific isotopic measurements by GC/IRMS. With the analyses of stable isotopes in environmental samples, from soils to sediment, water and plants we study:

  • Effects of changing environmental conditions on the mechanisms and rates of C stabilization in soils
  • Interactions between land degradation and C sequestration in soils
  • Reconstruction of past environments and climates to improve future predictions
  • Fate of organic pollutants in soils, sediments and water and interactions with organisms exposed to pollution
  • Application of CSIA in forensic science and food fraud

We have a RESPICOND respiration measurement facility to study transformations of organic matter and a glovebox for studies under anoxic conditions. In addition, concentrations of trace gases (CO2, CH4, N2O) can by quantified by GC.
Sample preparation facilities include a variety of extraction methods including Soxhlet, microwave, solid-phase (SPE) and accelerated solvent extraction (ASE), as well as a range of selective extraction methods for metals and organic substances.
Examples of research conducted in the laboratory include: i) the study of the molecular dynamics of organic matter transformations in soils to elucidate the role of soils in the global carbon cycle, ii) the presence and functioning of signaling chemicals in soils to study the interactions between above and belowground biodiversity, iii) the fate of anthropogenic substances in soil, sediment and water such as flame retardants, antibiotics, surfactants and polyfluorinated compounds, iv) biogeochemical cycling of elements (C, N, P) in soils and waters. The labs provide services to all three IBED themes and to colleagues from other Institutes from within and outside the UvA.

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Contact details

Eva de Rijke

Eva de Rijke, PhD
University of Amsterdam
Institute for Biodiversity and Ecosystem Dynamics

Visiting address
Sciencepark 904 (room C3.222)
1098 XH Amsterdam

email: E.deRijke@uva.nl

Andrea Gargano
Research

Smart Stationary Phases, Multidimensional Separations, Intact Proteins
Before moving to Amsterdam, I’ve been a fanciful mono-dimensional (separation) scientist for a number of years. In my PhD (Vienna, Austria) I worked at the synthesis and characterization of alternative stationary phase supports and chemistries (HILIC, mixed mode, ion exchange and chiral chromatography). My Post-Doctorate took me to Amsterdam at the HIMS institute and was my chance to learn and work in collaboration with several industrial partners (COAST project: HYPERformance LC) at the development and application of two-dimensional liquid chromatographic separations.

In 2015 I was awarded with a VENI grant by NWO for my research proposal on improving the separation technology for the characterization of Intact Protein Analysis (IPA). My research take place in the CASA laboratories (UvA and VU) and is performed in collaboration with international research laboratories (Pacific North West Laboratory, Richland WA, USA).

Contact details

gargano-andrea

Andrea Gargano, PhD
University of Amsterdam
Van 't Hoff Institute for Molecular Sciences (HIMS)

Postal address
Postbus 94157
1090 GD Amsterdam

Visiting address
Science Park 904 (room C2.248A)
1098 XH Amsterdam

telephone: +31 20 525 7040
email: a.gargano@uva.nl
researchgate.net/profile/Andrea_Gargano

Rob Haselberg
Research

I am an all-round analytical chemist with a strong background in capillary electrophoresis and liquid chromatography hyphenated with mass spectrometry. I use these technologies to characterize intact proteins. This ranges from proteins as small as insulin up to large protein complexes. I am interested in determining protein heterogeneity, getting insights in degradation processes, mapping intentional modifications and studying protein-protein interactions. Recently, my first personal research project was granted in which I will also focus on the bioanalysis of proteins. In all this research the goal is to keep the protein intact from sample to detector, as this reduces data complexity and minimizes risk of introducing artefacts along the way. As studying these properties of intact proteins is not straightforward, I develop and apply the required advanced hyphenated analytics. All of this is done in the context of biopharmaceutical, clinical, and doping analysis, often in collaboration with companies and other university groups. Besides my research, I also supervise the CE and CE-MS laboratory of the group.
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Contact details

haselberg-rob

Rob Haselberg, PhD
Vrije Universiteit Amsterdam
Amsterdam Institute for Molecules, Medicines and Systems

Postal address
de Boelelaan 1085
1081 HV Amsterdam

Visiting address
de Boelelaan 1108
1081 HZ Amsterdam

telephone: +31 20 598 7536
email: e r.haselberg@vu.nl

Suhas Nawada
Research
Contact details

Suhas Nawada, MSc
University of Amsterdam
Van 't Hoff Institute for Molecular Sciences (HIMS)

Postal address
Postbus 94157
1090 GD Amsterdam

Visiting address
Science Park 904 (room C2.248a)
1098 XH Amsterdam

email: S.H.Nawada@uva.nl