Project Details
Projekt
Directed evolution of limonene epoxide hydrolase to engineer its substrate- and enantioselectivity
Applicant
Professor Dr. Michael Arand
Subject Area
Biochemistry
Term
from 2004 to 2006
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 5427644
Epoxide hydrolases are increasingly important enzymes in biocatalysis for the production of chiral compounds. The goal of the project is to produce enzyme libraries that harbor variants of epoxide hydrolase with high activity and differential substrate selectivity in high frequency, using a semi-rational approach. The epoxide hydrolase gene will be synthesized using oligonucleotides that are degenerate in positions that encode the amino acids lining the substrate binding cavity of the enzyme, as deduced from the x-ray structure. These libraries will be screened for mutants with desired new properties, and the structural basis of these new properties will be analyzed by sequencing, biochemical characterization and 3D structure determination. Next, second- and third-generation libraries will be constructed, using the information obtained in the first analyses as a constraint (directed evolution). Close collaboration with the organic chemist Prof. Manfred Reetz (Director at the Max-PlanckInstitute for Carbon Research, Muelheim), who has developed exciting new approaches for the high throughput determination of enantioselectivities of enzymes and has filed a similar application using a complementary approach, is an integral part of the project.
DFG Programme
Priority Programmes
