Forschungs- & Entwicklungsinformationsdienst der Gemeinschaft - CORDIS

FP6

ENZYME MICROARRAYS Berichtzusammenfassung

Project ID: 25058
Gefördert unter: FP6-MOBILITY
Land: Germany

Final Activity Report Summary - ENZYME MICROARRAYS (Enzyme Microarrays-An integrated technology for the deconvolution of complex biochemical systems, drug discovery and diagnostics)

The project addressed complex research creating the tools for the deconvolution of complex biochemical processes. Activity-based inhibition of caspases, cathepsins, protein tyrosne phosphatase (PTPs), sentrin proteases (SENPs) was assessed both using classical solution and surface techniques by microarray technology. The enzymes of interest were cloned and purified so that collections of 10-20 different members of each enzyme class were available for the inhibition assays. New inhibitors - affinity labelling compounds - for each enzyme class were synthesised by classical organic and solid-phase peptide synthesis. The activity-based probes that were synthesized revealed to act as good irreversible inhibitors of the enzymes at very low concentrations both in solution and on-chip experiments.

A new route for the synthesis of the fluoromethylketones (as inhibitors of sentrin proteases) was developed, along with their incorporation into peptide mimics. In addition, new protein-tyrosine phosphatases inhibitors were designed and synthesized as small-molecules or incorporated into peptide skeletons and assayed as moderate to very good PTPs inhibitors. Besides study of enzyme inhibition using activity-based procedures, electronic detection of enzyme activity was approached in collaboration with other research group. This was based on constructing DNA-affinity labels conjugates which were finalised and assayed as basis for on-chip detection method of enzyme activity. Moreover, construction of supramolecular directed protein -arrays was a subject that was successfully finalised.

Different strategies were used to construct the supramolecular proteins arrays. Regardless the strategy involved, the construction of the metal-ion directed protein networks in a specific, directed and homogenous way required use of chemical proteomics techniques as methods to specifically modify proteins. This employed synthesis of affinity labels that in conjunction with polydentate ligands yielded affinity label conjugated ligands. Use of similar activity based probes in conjunction with methylmethacrylate yielded polymers that contain in their structure proteins positioned at precise locations, thus opening the way to a new field of protein-decorated polymers.

Kontakt

Johannes BUCHNER
Tel.: +49-89-289-13341
Fax: +49-89-289-13345
E-Mail-Adresse
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