Difference between revisions of "Part:BBa K2255005:Design"
 
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===Design Notes===
 
===Design Notes===
  
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D3 and the signal sequence are both the best conserved part from the attachment protein. So with protein global alignment (Needleman-Wunsch alignment), from two or three sequence at one time, we were eventually able to determinate D3 from M13.  
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The domain 3 and the signal sequence are both the best conserved part from the attachment protein. Using a global protein alignment (Needleman-Wunsch alignment), using two or three sequence at one time, we were eventually able to determinate D3 from M13.  
  
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We used iDT to optimise this sequence for ''E.coli''.
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This is the complete amino acid sequence of M13 protein 3:
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 +
<code>
 +
MKKLLFAIPLVVPFYSHSAETVESCLAKPHTENSFTNVWKDDKTLDRYANYEGCLWNATGVVVCTGDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGGTKPPEYG
 +
DTPIPGYTYINPLDGTYPPGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQGALTVYTGTVTQGTDPVKTYYQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDPFVCEYQ
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GQSSDLPQPPVNAGGGSGGGSGGGSEGGGSEGGGSEGGGSEGGGSGGGSGSGDFDYEKMANANKGAMTENADENALQSDAKGKLDSVATDYGAAIDGFIGDVSGLANGNGA
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TGDFAGSNSQMAQVGDGDNSPLMNNFRQYLPSLPQSVECRPFVFSAGKPYEFSIDCDKINLFRGVFAFLLYVATFMYVFSTFANILRNKES
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</code>
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We choose to only keep the D3 of this protein:
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 +
<code>
 +
AETVESCLAKSHTENSFTNVWKDDKTLDRYANYEGCLWNATGVVVCTGDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGGTKPPEYGDTPIPGYTYINPLDGTYP
 +
PGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQGALTVYTGTVTQGTDPVKTYYQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDLFVCEYQGQSSDLPQPPVNAGGG
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</code>
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We were able to found it because each domain is separated by a flexible sequence composed of Glycine and Serine <ref>Heilpern, A. J. & Waldor, M. K. pIIICTX, a predicted CTXphi minor coat protein, can expand the host range of coliphage fd to include ''Vibrio cholerae''. J. Bacteriol. 185, 1037–1044 (2003).</ref>. Then we retrotranslate this sequence in a nucleotidic sequence and we used iDT to optimise this sequence for ''E.coli'' production.
  
 
===Source===
 
===Source===

Latest revision as of 02:46, 2 November 2017


Domain 3 of p3 from M13 (Rfc25)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

The domain 3 and the signal sequence are both the best conserved part from the attachment protein. Using a global protein alignment (Needleman-Wunsch alignment), using two or three sequence at one time, we were eventually able to determinate D3 from M13.

This is the complete amino acid sequence of M13 protein 3:

MKKLLFAIPLVVPFYSHSAETVESCLAKPHTENSFTNVWKDDKTLDRYANYEGCLWNATGVVVCTGDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGGTKPPEYG DTPIPGYTYINPLDGTYPPGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQGALTVYTGTVTQGTDPVKTYYQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDPFVCEYQ GQSSDLPQPPVNAGGGSGGGSGGGSEGGGSEGGGSEGGGSEGGGSGGGSGSGDFDYEKMANANKGAMTENADENALQSDAKGKLDSVATDYGAAIDGFIGDVSGLANGNGA TGDFAGSNSQMAQVGDGDNSPLMNNFRQYLPSLPQSVECRPFVFSAGKPYEFSIDCDKINLFRGVFAFLLYVATFMYVFSTFANILRNKES

We choose to only keep the D3 of this protein:

AETVESCLAKSHTENSFTNVWKDDKTLDRYANYEGCLWNATGVVVCTGDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGGTKPPEYGDTPIPGYTYINPLDGTYP PGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQGALTVYTGTVTQGTDPVKTYYQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDLFVCEYQGQSSDLPQPPVNAGGG

We were able to found it because each domain is separated by a flexible sequence composed of Glycine and Serine [1]. Then we retrotranslate this sequence in a nucleotidic sequence and we used iDT to optimise this sequence for E.coli production.

Source

The initial sequence came from E.coli M13 filamentous phages. But we modified the sequence with iDT optimisation codon.

References

  1. ↑ Heilpern, A. J. & Waldor, M. K. pIIICTX, a predicted CTXphi minor coat protein, can expand the host range of coliphage fd to include Vibrio cholerae. J. Bacteriol. 185, 1037–1044 (2003).