Objective
TIM-barrel proteins are known to perform many different enzymatic functions. The active sites of these proteins are always formed by the 8 loops following after the 8 ß-strands. In this project we wish to study the structural determinants of the enzymatic activity, substrate specificity and stability of monomeric TIM-barrel proteins, in particular of monoTIM and two related chitinases. It is the aim of the project to understand how the TIM-barrel basic structure is employed by nature for different enzymatic activities. This understanding should in the end permit the use of the TIM-barrel scaffold for the design and construction of enzymes with desired properties. Several crystal structures of both enzymes (with or without ligand bound in the active site) are available. The monoTIM studies will be aimed at changing the specificity. By changing the sequences of loop-6, loop-7 and loop-8 it will be attempted to make a sugar binding pocket (instead of a phosphate binding pocket) such that the specificity will be converted from a triosephosphate isomerase into a xylose isomerase, which is an important industrial enzyme in the food industry. The chitinase studies will be aimed at making a more simple, more active and more stable chitinase with a rationally designed substrate specificity. For the latter purpose comparative studies of the related chitinases will be conducted and site directed mutagenesis will be used to verify the role of their differences. Mutagenesis work on the chitinases will further concern for example replacing long protruding loops by short connections as well as making designed changes in active site loops. New chitinase variants will have important industrial applications in plant biotechnology. The expertise essential for these projects is abundantly available in this project, in particular protein crystallography, modelling, genetic engineering, biochemistry and enzymology, as well as expertise in organic chemistry and calorimetry which is essential for making special substrates and to study stability and binding constants, respectively. The major thrust of the project is to characterise chitinases and monoTIM further, as well as to rationally design new functional properties, in particular via changing the loops. It is anticipated that new rules will emerge from these studies that are of crucial importance for the redesign of protein function.
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.
- natural sciences chemical sciences organic chemistry
- natural sciences earth and related environmental sciences geology mineralogy crystallography
- medical and health sciences medical biotechnology genetic engineering
- natural sciences chemical sciences analytical chemistry calorimetry
- natural sciences biological sciences biochemistry biomolecules proteins enzymes
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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
69012 HEIDELBERG
Germany
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