Part:BBa_K320006

Constitutive expression of Immunotoxin

This BioBrick allows the expression of the Immunotoxin ( BBa_K320001) regulated by a strong constitutive promoter (BBa_K320002).

The Immunotoxin BioBrick (BBa_K320001) is composed of two main parts. The first is a single-chain antibody fragment (scFv) directed to Pbs2l, a surface membrane protein of Plasmodium berghei ookinetes (sexual reproductive stage of a close relative of the Malaria parasite P. falciparum). This fragment is linked to the second part, the lytic peptide Shiva-1. Shiva-1 is a synthetic peptide, whose amino acid sequence is similar in length and amino acid properties to Cecropin B (38 amino acids), an antibacterial peptide. The purpose of the Immunotoxin is to specifically target and lyse the ookinete.

Previous experiments showed that feeding mosquitoes with recombinant E. coli expressing this Immunotoxin induced a significantly lower oocyst density. [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6T29-42JHDJD-9&_user=10&_coverDate=03%2F31%2F2001&_rdoc=1&_fmt=high&_orig=search&_origin=search&_sort=d&_docanchor=&view=c&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=5f4b78b08a1846e241faaed33bc76cb3&searchtype=a reference]

However E. coli is not a natural symbiont of the mosquito. We aim to improve this experiment by making Asaia express the immunotoxin in the mosquitoes intestinal tract, and run experiments to see if it significantly reduces the number of oocysts.

This BioBrick (BBa_K320001)also contains an export signal sequence to export the immunotoxine protein to the periplasm.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 62
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 883
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]