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Recombinant expression and in vitro folding of a Class A GPCRs
Germany (bvmd) - Universitaet Leipzig, Leipzig
Biophysics and Medical Physics
Institute of Medical Physics and Biophysics, Leipzig
Prof. Dr. Daniel Huster
Type of Research Project
- Basic science
What is the background of the project?
The candidate will be involved in the recombinant expression of the Y2 receptor in E. coli by high cell density fermentation and in cell free expression. Under these conditions, the receptor is produced in non-native aggregated form, but can be expressed either uniformly of specifically isotopically labeled. The labeled aggregates will be isolated, solubilized and purified in a two-step purification using the FPLC-system (size exclusion and affinity chromatography). The student will perform subsequently refolding of the receptor into different phospholipid environment in functional form. Finally the samples have to be concentrated for NMR-Measurements.
What is the aim of the project?
The aim of the project is the investigation of the domain-specific dynamic of the Y2 receptor in different time-scales using various methods of solid-state NMR.
What techniques and methods are used?
What is the role of the student?
What are the tasks expected to be accomplished by the student?
Will there be any theoretical teaching provided (preliminary readings, lectures, courses, seminars etc)
What is expected from the student at the end of the research exchange? What will be the general outcome of the student?
What skills are required of the student? Is there any special knowledge or a certain level of studies needed?
FLUENT English is absolutely necessary! Preliminary reading will be helpful for better understanding of experiments; there will be also discussions and individual seminars, furthermore the student will attend at a postgraduate seminar and the institute colloquium (both weekly).
Are there any legal limitations in the student’s involvement
Type of students accepted
This project accepts: - Medical students
- Biotechnol Prog. 2009 Nov-Dec;25(6):1732-9.;Prokaryotic expression; in vitro folding; and molecular pharmacological characterization of the neuropeptide Y receptor type 2; Schmidt P et al
- A reconstitution protocol for the in vitro folded human G protein-coupled Y2 receptor into lipid environment; Schmidt P et al
- Oxidative in vitro folding of a cysteine deficient variant of the G protein-coupled neuropeptide Y receptor type 2 improves stability at high concentration
- The G-protein-coupled neuropeptide Y receptor type 2 is highly dynamic in lipid membranes as revealed by solid-state NMR spectroscopy.; Prokaryotic expression; in vitro folding; and molecular pharmacological characterization of the neuro
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