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Workgroup Protein Interactions: Investigators of enzymatic activity in proteins

The Workgroup Protein Interactions at the Ruhr-Universität Bochum investigates mechanisms in the interactions and enzymatic activity of proteins involved in immunity and disease

Human guanylate binding proteins from the family of large GTPases, as well as small GTP binding proteins like the oncogene Ras, employ nucleotide binding and hydrolysis in protein interactions in order to switch between different structural states and to alter the functional status.

Biochemical and biophysical methods help to understand the interplay of these proteins on the molecular level and to relate quantitative characteristics of protein interactions to the biological activity.

Protein interactions: the key agents in living organisms

The interactions of proteins with each other, and with other molecules, as well as with water as solvent and other cosolvents, form the basis of most biological processes. Using biochemical and biophysical techniques, as well as self-developed methods, Workgroup Protein Interactions investigates the interactions between proteins that play an important role in the mediation of cellular signals.

In addition, the Workgroup Protein Interactions investigates molecular processes in the enzymatically catalysed hydrolysis of guanosine triphosphate (GTP), which is important for the control of protein interactions.

On the one hand, our work aims to elucidate the specific mechanisms of action of the examined proteins and, on the other hand, to derive universally valid laws.


In order to study the interactions of proteins of interest, Workgroup Protein Interactions first needs to prepare and purify them. Using a suitable expression vector, the respective protein is synthesised in bacterial cultures and then purified by different chromatographic methods in two or three steps.

Standard molecular biological methods (in particular PCR, polymerase chain reaction) allow the construction of desired variants of the proteins. In particular, the introduction of certain point mutations or the deletion of one or more protein domains, followed by the biophysical studies of the altered properties and the comparison with the wild type allow meaningful conclusions on the functioning of the proteins.

  • Thermodynamic characterization of the formation of protein complexes;
  • Stability of proteins in solutions with ionic liquids;
  • More complex Ras/effector interactions;
  • Kinetic analysis of protein interactions;
  • Influence of macromolecular cosolvents on the formation of protein complexes;
  • Structural study of protein complexes;
  • Mechanisms of enzymatically catalyzed GTP hydrolysis;
  • Self-assembly of large GTP-binding proteins;
  • Identification of cellular interaction partners and analysis of protein contact surfaces; and
  • Determination of contact surfaces in protein complexes and mutation analysis.

Main topics include:

  • Molecular mechanisms of human Guanylate Binding Proteins (hGBP):
    • The enzymatic activity of hGBP leading to successive cleavage of the gamma- and the beta-phosphate uniquely facilitates a cascade of structural changes;
    • The extent and the dynamics of intramolecular domain movements, as investigated with the help of site specific fluorescence labels, controls the interaction with other hGBP isoforms; and
    • Nucleotide binding and hydrolysis is responsible for i) the anchorage of hGBP at different endo-membranes, ii) hGBP self-assembly and the formation of polymers, iii) the powering of a cycle of biological activity.
  • Interactions between Ras and effector proteins:
    • Thermodynamic and transient kinetic analysis of Ras/effector interactions reveal mechanisms of interaction and mutual control of activity;
    • Expanding this on mutational analyses yields a quantitative understanding of interaction specificity and molecular recognition; and
    • The malfunction of oncogenes like Ras and of tumour suppressor genes like Nore1/RASSF5 is directly connected to changes in enzymatic activity and binding affinities, respectively, evoked by point or deletion mutations.
  • The role of water solvation in protein interactions and stability:
    • The stability of proteins in terms of unfolding and of aggregation is critically dependent on the properties of the solvent, i.e. water together with many possible co-solvents and co-solutes; and
    • The aim of thermodynamic studies with the help of isothermal titration calorimetry, ITC, and differential scanning calorimetry, DSC, is a profound understanding of the impact of water perturbations on the stabilising or destabilising interactions with proteins.

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Workgroup Protein Interactions on immune defence

The Faculty of Chemistry and Biochemistry at Ruhr-University Bochum, Germany, unites the highest level of research and innovative teaching. Focusing on interfacial systems chemistry and...

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