Objective
- To understand the mechanisms of wheat gluten protein systhesis, transport, folding and assembly in relation to the formation of protein bodies and the structure and functional properties of gluten.
Results so far
- The production of suitable levels of in vitro synthesised gamma-gliadin in a cell free system; this has been facilitated by cloning the coding sequence of the gamma-gliadin cDNA into the pGEM3 transcription vector. Subsequent transcription and translation using this template resulted in easily detectable levels of radiolabelled protein.
- The gamma-gliadin pGEM template has been used to set up protein translocation assays in a cell free system containing pancreatic canine microsomal membranes. The translocation of the gamma-gliadin was observed, as determined by signal peptide cleavage and protease-protection assays. The use of stalled translocation-intermediates was attempted to examine the interaction between the gamma-gliadin-translocating nascent chain and chaperones such as BiP and PDI. Initial attempts utilised long nascent chains of greater than 20 kD in length. However these polypeptides have failed to produce crosslinks, indicating that such protein-protein interactions occur when the nascent chain initially enters the ER lumen. Therefore, shorter nascent chains have been generated and are at present being examined in vitro.
- Transgenic tobacco lines expressing gamma-gliadins with modified targeting information are being used for electron microscopic studies. The ultrastructure (in particular of the endomembrane system) is being examined. Immunogold labelling is being optimised for this particular system to allow the precise determination of the subcellular locations of the transgene products.
- Expression of an epitope-tagged version of the gamma-gliadin in transgenic wheat is also being carried out to determine the deposition of the protein in a homologous background. The c-myc epitope tag has been introduced, in frame, into the C-terminus of the gamma-gliadin, and this will be expressed in wheat under the control of the HMW-glutenin promoter. Transgene expression will be detected by the use of the 9E10 monoclonal against the c-myc tag.
- To determine the pathway and mechanism of gamma-gliadin translocation across the ER membrane, including interactions with proteins of the translocation machinery;
- To determine the stage at which the chaperone BiP interacts with the nascent gamma-gliadin chain to aid protein folding;
- To determine the effects of ER-localised enzymes and molecular chaperones on the folding of wheat storage proteins and their assembly into protein bodies using an in vitro system with proteins expressed in E. coli;
- To determine the assembly into protein bodies of natural and modified storage proteins in transgenic tobacco plants, and the effects of over-expression of ER localised enzymes and molecular chaperones on these processes.
Fields of science (EuroSciVoc)
CORDIS classifies projects with EuroSciVoc, a multilingual taxonomy of fields of science, through a semi-automatic process based on NLP techniques. See: The European Science Vocabulary.
CORDIS classifies projects with EuroSciVoc, a multilingual taxonomy of fields of science, through a semi-automatic process based on NLP techniques. See: The European Science Vocabulary.
This project has not yet been classified with EuroSciVoc.
Be the first one to suggest relevant scientific fields and help us improve our classification service
You need to log in or register to use this function
Programme(s)
Multi-annual funding programmes that define the EU’s priorities for research and innovation.
Multi-annual funding programmes that define the EU’s priorities for research and innovation.
Topic(s)
Calls for proposals are divided into topics. A topic defines a specific subject or area for which applicants can submit proposals. The description of a topic comprises its specific scope and the expected impact of the funded project.
Data not available
Calls for proposals are divided into topics. A topic defines a specific subject or area for which applicants can submit proposals. The description of a topic comprises its specific scope and the expected impact of the funded project.
Call for proposal
Procedure for inviting applicants to submit project proposals, with the aim of receiving EU funding.
Data not available
Procedure for inviting applicants to submit project proposals, with the aim of receiving EU funding.
Funding Scheme
Funding scheme (or “Type of Action”) inside a programme with common features. It specifies: the scope of what is funded; the reimbursement rate; specific evaluation criteria to qualify for funding; and the use of simplified forms of costs like lump sums.
Funding scheme (or “Type of Action”) inside a programme with common features. It specifies: the scope of what is funded; the reimbursement rate; specific evaluation criteria to qualify for funding; and the use of simplified forms of costs like lump sums.
Coordinator
BS41 9AF BRISTOL
United Kingdom
The total costs incurred by this organisation to participate in the project, including direct and indirect costs. This amount is a subset of the overall project budget.