Objective
This concerted effort by academic and industrial laboratories aims at improved understanding of structure-function relationships in enzymes synthesizing or degrading commercially valuable alpha-glucans. The goals are to tailor-make specificities i) in glucosyltransferases utilising sucrose for production of new prebiotic oligo- and poly-alpha-glucans to be applied in cosmetics, food, and feeds by BioEurope (Partner and SME expert in enzymic sugar synthesis) and ii) in hydrolases with novel or improved activities specific for alpha-1,4 and 1,6-bonds in starch, and branched, linear or cyclic oligosaccharides, thus addressing a need for multi-specific amylases, an ambitious target crucial in efficient starch utilization. To develop engineered hydrolases we collaborate with Carlsberg Research Laboratory on transgenic plants and Consortortium fur elektrochemische Industrie on enzymic environment- friendly industrial waste cleaning and sugar synthesis.
Scientifically, the project is an integrated, multi-disciplinary approach, combining 3D structure determination, design and production of enzyme variants. The target enzymes bacterial glucosyltransferases, barley alpha-amylase and limit dextrinase, and bacterial cyclodextrinase are unified by two properties, i) a common catalytic alpha/beta-barrel domain, which we discovered in glucosyltransferases in a novel circularly permuted form and ii) structurally related substrates and products. We will provide evidence on enzyme-substrate interactions that describe new specificities at the atomic level and enable development of designer molecules by rational protein engineering. The project conforms to the area 6. Structural Biology, 6.1 Structure-function relationship with emphasis an development of biomolecules with desired functions, and will address:
1. Delivered glucosyltransferase and starch/oligosaccharide hydrolase variant for exploitation of industrial potential as outlined above.
2. Crystal structure determination of enzymes with new and engineered specificities, isolated domains, novel domain fusions and multi-domain complexes or enzymes stabilized by bifunctional inhibitors. Time-resolved and molecular dynamics studies of the barley alpha-amylase mechanism.
3. 1D/3D structure comparisons as basis i) for localised random mutagenesis in binding regions/sequence motifs coupled with screening for altered specificity, and ii) for rational design of variants with putative substrate/ product specificity modification. Combination of functionalities in wild-type and engineered fusions with the starch binding domain to enhance attack on solid starches as monitored by atomic force microscopy, used also in mutational discrimination of the two domain binding sites.
4. Synthesis of new non-convertible thio-glucoside analogs and photoactivatable oligosaccharides used in static and time-resolved crystallography, enzyme kinetics and binding thermodynamics analyses.
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 earth and related environmental sciences geology mineralogy crystallography
- natural sciences physical sciences thermodynamics
- natural sciences physical sciences optics microscopy
- natural sciences biological sciences biochemistry biomolecules proteins enzymes
- natural sciences biological sciences molecular biology structural biology
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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
2500 VALBY
Denmark
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