PROFACT · Protein factory for kinase substrate searching and protein detecting micro-array development
FP6 — Marie Curie Actions (Human Resources and Mobility)
- Duration
- 2005-01-01 → 2005-12-31
- EU contribution
- €37,113
- Participants
- 1
- Scheme
- ERG
Lines connect the coordinator with its partners.
Results in brief
Final Activity Report Summary - PROFACT (Protein factory for kinase substrate searching and protein detecting microarray development)
In biological terms, the 21st century is the era of postgenomics research facing the challenging task of untangling and revealing the function of subtle protein networks of living organisms. The large scale studies urge invention of high-throughput approaches capable coping with analysis of thousands of proteins. This new field of biology -the proteomics- largely depends on availability of ample amount of a large number of proteins. According to the objectives of the proposal, we set up a wheat germ extract based cell free protein translation system that meets the requirements of proteomics by producing proteins of interest rapidly and efficiently. The established protein translation system was applied to produce active protein kinases for identification of their putative substrates. We aim at isolation of target molecules of protein kinases located in mitochondria, since the mitochondria is central cell organ not just as main energy converter but also as central signal integrator. Revealing protein phosphorylation pathways in mitochondria will contribute to understanding of regulation of programmed cell death, an important procedure in development and eradicating tumour cells. In order to implement this task plant mitochondrial protein extracts have been purified and the produced protein kinase will be added to these extracts to label the target molecules. An equally important objective of the proposal is the isolation of aptamers against proteins. Aptamers are short oligonucleotides binding the target proteins and macromolecules with high specificity and affinity, thus, rivals of antibodies in many applications like, protein and macromolecule detection, affinity purification, immuno-affinity labelling and even used as human therapeutics. In our laboratory, a random aptamer library was designed and used for identification of specific oligonucleotides for protein kinases and mycotoxins. In both cases, these works resulted in isolation of aptamers binding to the target molecules, and possible applications of these oligonucleotides are currently tested.
Data: CORDIS, © European Union
Project objective
After decoding the genomes of several living organisms, the main interest of biologists is focused on protein studies. This new field of biology -the proteomics- is signified by large-scale protein analysis and needs high-throughput approaches. Progress of the proteomics studies are largely depends on availability of sufficient amount of a large number of proteins. Presently, the protein production is cumbersome and has to be optimised on case-by-case basis. We aim at delivering solution for this bottleneck of proteomics by devising a 'protein factory'. Elimination this rate-limiting step speeds up the proteomics research. We exploit the 'protein factory' in two applications, first, to devise a general substrate hunting protocol; second to invent protein-detecting micro-arrays (PDM). Identification of protein kinase substrates is a major challenge of the post-genomic era and requires improved methodologies. Despite several efforts with different methods the identification has been slow and intricate.We describe a feasible protocol suitable for high-throughput substrate screening in any organism. We use this protocol for identification of plant kinase substrates involved in pathogen responses; hence, the expected results help understanding this agriculturally important process. PDMs are the protein sensors of future and could be used -besides basic science- in human diagnostic, food industry and environmental sciences. Development of these chips requires ample amount of assorted proteins, which could be conveniently produced with our system.During my training, I participated in European and national projects and established essential connections. The complexity of research objectives needs dedicated experts from a number of field therefore urge cross-border and national co-operations. I can transfer my acquired knowledge to university offering a position in a multidisciplinary environment.
Original text from CORDIS.
Participants
- BUDAPEST UNIVERSITY OF TECHNOLOGY AND ECONOMICS · BUDAPESTCoordinatorHungary
Links
Data: CORDIS, © European Union