Difference between revisions of "Part:BBa K2324008"
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The literature has shown that the terminal pili protein FimH (Le Trong et al 2010) can be modified by inserting heterologous sequences at position 225 and 258 (Pallesen et al 1995, Shembri et al 1999). However these two amino acids are in the pilin binding domain which may present difficulties when attempting to introduce large modifications. Harvard iGEM 2015 also introduced modifications at position 258 of the mature FimH protein.</p>
 
The literature has shown that the terminal pili protein FimH (Le Trong et al 2010) can be modified by inserting heterologous sequences at position 225 and 258 (Pallesen et al 1995, Shembri et al 1999). However these two amino acids are in the pilin binding domain which may present difficulties when attempting to introduce large modifications. Harvard iGEM 2015 also introduced modifications at position 258 of the mature FimH protein.</p>
 
<p>
 
<p>
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This part produces a FimH adhesin protein fused with GFP at its 258th amino acid residue. The coding sequence is under the control of a rhamnose-inducible promoter, with a B0034 RBS and a B0015 terminator. The part, when induced, produces a fluorescent FimH protein that should initiate pilus biosynthesis when co-transformed with a plasmid containing the <i>fim operon</i></p>.
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This part produces a FimH adhesin protein fused with sfGFP at its 258th amino acid residue after signal peptide cleavage. The coding sequence is under the control of a rhamnose-inducible promoter, with a B0034 RBS and a B0015 terminator. The part, when induced, should produce a fluorescent FimH protein that should initiate pilus biosynthesis when co-transformed with a plasmid containing the <i>fim operon</i></p>. Unfortunately despite best efforts (multiple growth temperatures and multiple rhamnose inducing concentrations) no fluorescence has been detected in any of the <i>E. coli</i> strains.
  
 
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Revision as of 21:47, 1 November 2017


pRha_FimH_258sfGFP

The literature has shown that the terminal pili protein FimH (Le Trong et al 2010) can be modified by inserting heterologous sequences at position 225 and 258 (Pallesen et al 1995, Shembri et al 1999). However these two amino acids are in the pilin binding domain which may present difficulties when attempting to introduce large modifications. Harvard iGEM 2015 also introduced modifications at position 258 of the mature FimH protein.

This part produces a FimH adhesin protein fused with sfGFP at its 258th amino acid residue after signal peptide cleavage. The coding sequence is under the control of a rhamnose-inducible promoter, with a B0034 RBS and a B0015 terminator. The part, when induced, should produce a fluorescent FimH protein that should initiate pilus biosynthesis when co-transformed with a plasmid containing the fim operon

. Unfortunately despite best efforts (multiple growth temperatures and multiple rhamnose inducing concentrations) no fluorescence has been detected in any of the E. coli strains.

Sequence and Features


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]