Prof. Dr. Volker Dötsch (Frankfurt): Combination of cell-free protein expression and liquid state NMR spectroscopy for the structure determination of membrane proteins |
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| Startdatum/-zeit: | 12.06.2012 17:15 |
| Enddatum/-zeit: | 12.06.2012 |
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| Adresse: | SFB 944, Prof. Dr. Ch. Ungermann, Prof. Dr. Jacob Piehler apl. Prof. Dr. Merzendorfer Barbarastr. 11 49076 Osnabrück |
| Telefon: | 0541/969-3502 |
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| E-Mail: | merzendorfer@biologie.uni-osnabrueck.de |
| Homepage: | http://www.biologie.uni-osnabrueck.de/Fachbereich/?x=aaad,aczx,gn |
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| Adresse: | Biologiegebäude, Hörsaal 35/E01 Barbarastr. 11 49076 Osnabrück |
| Karte: | Bei Google Maps anzeigen |
| Beschreibung: | Cell-free expression systems are ideal for the production of proteins that cannot be overexpressed in E. coli cells. In particular for NMR spectroscopy, cell-free expression provides major advantages since it allows for amino acid type selective labeling without metabolic scrambling. We have developed several labeling protocols that allow us to obtain the backbone assignment of membrane proteins in an efficient way. The TMS-labeling (Transmembrane segment enhanced labeling) is based on the fact that 60% of the amino acids of the transmembrane helices consist of only six different amino acid types. Double labeling membrane proteins with these six amino acids produces large consecutive stretches of labeled amino acids that can be analyzed with standard triple resonance experiments with a significantly reduced peak overlap. In addition, we have developed a combinatorial labeling scheme that allows us to assign specific amino acids. Using these methods we have obtained the backbone assignment for the C-terminal fragment of presenilin, the catalytic component of the -secretase which is responsible for cleaving the amyloid precursor protein. This peptide forms the plaques that are found in the brains of Alzheimer patients. We have determined its structure as well as the structure of the retinal binding protein Proteorhodopsin. In addition, we have studied the interaction between the soluble Get3 dimer and its membrane bound receptors Get1 and Get2 and determined the crystal structures of both complexes. The entire Get complex is responsible for the membrane insertion of tail-anchored membrane proteins. |
| Kategorie: | SFB-Seminare (Biologie) |
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