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Homology search of Defensins among different species

Homology search of Defensins among different species. HT05 Fall Research Training Project of MBB Program XIA Han. The Purpose of My Project. Database of Defensins 10~40 amino acids length OS: Plant, Invertebrate and Vertebrate Literatures related to this sequences

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Homology search of Defensins among different species

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  1. Homology search of Defensins among different species HT05 Fall Research Training Project of MBB Program XIA Han

  2. The Purpose of My Project • Database of Defensins • 10~40 amino acids length • OS: Plant, Invertebrate and Vertebrate • Literatures related to this sequences • Search for conversed sequence patterns • Connect sequence’s features with structure

  3. What is DEFENSIN? • Antimicrobial Peptide (AMP) • Germline-encoded • Widespread • Cysteine-rich • Wide antimicrobial spectrum

  4. Structural Features • 4~8 cysteine (mostly) • 2~4 disulfide bridges • Lineal or cyclic molecule • Cationic • Amphipathic Tang, Y. Q. et al. Science 286, 498-502 (1999)

  5. Construction of Database • Stage 1: Search for sequences • Protocol for search • Literature Search • Database Search • Data Refine and Validation

  6. Database Search • AMSDb Antimicrobial Sequences Database http://www.bbcm.units.it/~tossi/amsdb.html • APD Antimicrobial Peptide Database http://aps.unmc.edu/AP/main.php • NCBI

  7. Data Refinement and Validation • Refinement Strategy • 10~40 amino acids • 2~4 disulfide bridges • Validation Strategy • Consistent with corresponding entry in NCBI

  8. Construction of Database • Stage 2: Accession of Data • Classification : Based on Taxonomy • Format : FASTA • Additional information : References of Sequences

  9. Summary of Database

  10. Sequence Alignment • ClustalX • Multiple alignment && Profile alignment • Alignment based on evolutionary tree • More interactive • More Protein Weight Matrices • BLOSUM (Henikoff) • PAM (Dayhoff) • GONNET

  11. Test Runs • Parameters • Default settings • GONNET series as Protein Weight Matrices • Pairwise alignment: GONNET 250 • Multiple Alignment: GONNET series However……

  12. Test Runs Results

  13. Test Runs Results

  14. Profile Alignment • Important Parameters • End Gap Separation : On • Residue Specific Penalties and Hydrophilic Gap Penalties : On • New Gaps before Alignment and Reset All Gaps before Alignment : Off • BLOSUM series as Protein Weight Matrices • Pairwise alignment: BLOSUM 30 • Multiple Alignment: BLOSUM series

  15. Profile Alignment • Profile Alignment in a designed order : • Profile alignment multiple-aligned insect sequences with multiple-aligned Arachnida sequences, get P1.aln (26 sequences) ; • P1.aln with multiple-aligned Mussel sequences, get P2.aln (29 sequences) • P2.aln with multiple-aligned vertebrate sequences, get P3.aln (106 sequences) • P2.aln with multiple-aligned plant sequences, get P4.aln (40 sequences)

  16. Profile Alignment Result: P2.aln

  17. Structural Alignment • Secondary Structural Prediction JPRED(http://www.compbio.dundee.ac.uk/~www-jpred/ ) • Sequence-based Structural Alignment • BLAST • MSDlite(http://www.ebi.ac.uk/msd-srv/msdlite/index.html)

  18. Profile Alignment Result: P2.aln

  19. Superposition : 1L4V and 1ICA

  20. Acknowledgements: • My Supervisor : Anders Backlund • Everyone in Department of Pharmacognosy • Margareta Krabble • Mikael Thollesson • My Supporting, lovely Classmates!

  21. References • Bulet, P., Stocklin, R. & Menin, L. Anti-microbial peptides: from invertebrates to vertebrates. Immunol Rev 198, 169-84 (2004). • Tang, Y. Q. et al. A cyclic antimicrobial peptide produced in primate leukocytes by the ligation of two truncated alpha-defensins. Science 286, 498-502 (1999). • Gueguen, Y. et al. Characterization of a defensin from the oyster crassostrea gigas: Recombinant production, folding, solution structure, antimicrobial activities and gene expression. J Biol Chem (2005). • Hoffmann, J. A., Kafatos, F. C., Janeway, C. A. & Ezekowitz, R. A. Phylogenetic perspectives in innate immunity. Science 284, 1313-8 (1999). • Mygind, P. H. et al. Plectasin is a peptide antibiotic with therapeutic potential from a saprophytic fungus. Nature 437, 975-80 (2005).

  22. Tack sä mycket!God Jul och Gott Nytt Är!

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