Membrane protein session Erice

Introduction Werner Kühlbrandt5 min Aquaporin 0Tom Walz30+5 min Na + /H + antiporterCarola Hunte30+5 min EM and x-rayWerner Kühlbrandt30+5 min Coffee break30 min Photosystem II Jan Kern30+5 min Solid-state NMRHartmut Oschkinat30+5 min Programme

in pdb deposits

all PDB entries , membrane protein entries

What makes it so hard ? Few membrane proteins are naturally abundant Most are difficult to express Difficult to purify Difficult to crystallize Difficult to solve

Usual problems Insufficient protein available Protein unstable in detergent solution Protein precipitates upon concentration Detergent interferes with crystal order Bound lipids may or may not be required Crystals are often small, poorly ordered and anisotropic Crystals often cannot be frozen

Different types of membrane protein crystals Type 1 crystalsType 2 crystals “Type 4” crystals “Type 3” crystals

Use of membrane protein crystals 3D crystals for X-ray crystallography –Protein in detergent solution –Initial crystals often poorly ordered –Antibody fragments as spacers –Lipidic cubic phases ? 2D crystals for electron microscopy –Protein in lipid bilayer –Physiological conditions –Good probability –Experimental phases Small crystals for solid-state NMR

Techniques for structure determination X-ray crystallography (Carola Hunte, Jan Kern) –atomic coordinates –single state (unless you are lucky) Electron crystallography –2D crystals: 3 Å - 8 Å resolution with ab initio phases (1.8 Å by electron diffraction: Tom Walz) –protein in membrane –different conformational states (Werner Kühlbrandt) Single-particle EM –6 Å - 20 Å resolution –protein in solution or in membrane NMR –Solution: small  -barrels –Solid-state (Hartmut Oschkinat)