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On the evolution of membrane proteins Homology detection Blast, PSIBLAST are they good ? SHRIMP (Bernsel, submitted) Evolution of TM-helices Internal duplications >50% of +6TM proteins Arne Elofsson ([email protected]) Problems with homology detection of TM proteins Methods optimized on globular proteins High scoring false positives due to similarity between unrelated TM regions Hedman et al, 1998 (Pmembr): Fold recognition methods improve this fact Globular detections improved by profileprofile (HMM-HMM) comparisons Arne Elofsson ([email protected]) Profile-profile methods improve detection of membrane proteins Arne Elofsson ([email protected]) Classification of GPCRs at 1% FPR Arne Elofsson ([email protected]) How does TM proteins evolve Mutation rates any differences in respect to the different compartments. Few “multi-domain” membrane proteins Many complexes Internal symmetry Arne Elofsson ([email protected]) Slower evolution in the membrane Arne Elofsson ([email protected]) Internal duplications on sequence level Searched for internal duplication events in 87 prokaryotic genomes Out of 38,124 membrane proteins, 377 duplication events could be detected (~1%) Internal duplications has been hard to detect on sequence level α α X J. Mol. Biol. 2004, 339:1-15 α α A α B Arne Elofsson ([email protected]) α C Lactose Permease Arne Elofsson ([email protected]) 1xqf (NH3-channel) Antiparallel duplication Arne Elofsson ([email protected]) 12-TM chain from Cytochrome C-oxidase(Triplication) Arne Elofsson ([email protected]) Internal symmetry in α-helical inner membrane proteins N Antiparallel: LeuT 2-fold symmetry axis SecY BtuCD Parallel to membrane CAQP1 GlpF Odd number of TMH AmtB ClC H+/Cl- exchange transporter Domains anti-parallel to NhaA each other middle C Parallel: LacY GlpT N Sav1866 ABC transporter AcrB EmrD ADP/ATP carrier In plane with membrane Even number of TMH Domains parallel to each other Arne Elofsson ([email protected]) Many (most) proteins have internal duplications Fraction of chains with internal duplications Fraction containing a duplication 1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 6 8 10 Number of transmembrane helices (minimum) Arne Elofsson ([email protected]) 12 Duplications in TM proteins Distribution of duplication size 100,00% 90,00% 80,00% 70,00% 60,00% 50,00% 40,00% 30,00% 20,00% 10,00% 0,00% No duplic ation 6 T MH 5 T MH 4 T MH 3 T MH 6 7 8 9 10 11 ³12 Number of transmembrane helices in chain Arne Elofsson ([email protected]) Both parallel and antiparallel duplications exist Parallel / Antiparallel duplications 6 5 4 Antiparallel 3 Parallel 2 1 0 3 4 5 Number of duplicated helices Arne Elofsson ([email protected]) 6 But in order to have antiparallel domains, they must be inserted with opposing topology! C N inside membrane • outside C In order for an antiparallel symmetry to be explained by a duplication event, each subunit of the homodimer must be inserted with opposite topology • That implies that a sequence can have dual topology, be inserted either way in the membrane • Have been constructed in labs (Cell 62, 1135-1141 & Cell 77, 401412) N Arne Elofsson ([email protected]) Hypothesis for antiparallel internal symmetry Nat. Struct. Mol. Biol. 2006, 13: 94-96 Arne Elofsson ([email protected]) Detection of internal duplications Structural alignments Sequence alignments Finds most of the manually detected examples. Sequence alignments do not find most. Using SHRIMP about 80% can be found Also finds a few extra. Some problems defining exact duplication unit Genome scanning Ongoing. Arne Elofsson ([email protected]) Acknowledgements Erik Granseth Anni Kauko Jenny Falk Andreas Bernsel Kristoffer Illergård Arne Elofsson ([email protected]) Final comments Thanks to EMBRACE Please fill out the questionaire.. Arne Elofsson ([email protected])