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Team:Paris/Addressing overview2
From 2009.igem.org
iGEM > Paris > Adressing > ClyA
Introduction
Bacterial pathogens display proteins on their surface that may interact with their hosts in order to mount successful infections. Although the primary function of the peptidoglycan is to provide a physical barrier for protection against both mechanical and osmotic stresses, it also serves as a scaffold to anchor external structures such as the outer cell membrane in Escherichia Coli. Over the past 20 years, it has become apparent that Gram-negative bacteria have evolved a variety of mechanism by which proteins are displayed on the cell surface (Tat and Sec transporter for example, cf the section "to the periplasm, export system" for more information about them). Blablabla One of the major protein of the bacterial membrane is OmpA.
There is also an other protein but less known than OmpA which can exported from cytoplasm to outer membrane then by vesiculation interact with their host.
OmpA :
ClyA :
Pore-forming toxins (PFTs) are a class of potent virulence factors that convert from a soluble form to a membrane-integrated pore. They exhibit their toxic effect either by destruction of the membrane permeability barrier or by delivery of toxic components through the pores.
Cytolysin A (ClyA, also known as HlyE), a PFT, is a cytolytic α-helical toxin responsible for the haemolytic phenotype of several Escherichia coli. (1)
Why did we decided to use ClyA?
Althought the toxic effect of Cly A, using it is an interesting way to adress protein to the external membrane, because ClyA contain the signal peptide required to be exported form the cytoplasm to the outer membrane. In addition, it has been shown that certain membrane and/or soluble periplasmic proteins are enriched in vesicles while others are preferentially excluded. The majority of these enriched proteins happen to be virulence factors including cytolysin A of Escherichia Coli.(2)
Previous studies demonstrated that genetic fusions between E. coli ClyA and reporter proteins such as Bla and GFP were translocated across the cytoplasmic membrane (3,4) and that localization was independent of the position (N or C terminus) of ClyA in the fusion protein. Moreover a recent article (5) demonstrated that fusions to the C terminus of ClyA allow the transport of protein closer to the surface of outer membrane and to extend them into the extracellular environment.
Bibliography :
(1) M.Mueller, U.Grauschopf, T.Maier, R.Glockshuber1 & N.Ban, The structure of a cytolytic a-helical toxin pore reveals its assembly mechanism, 2009, Nature vol 459 , 726-731
(2)S.N. Wai, B.Lindmark, T.Soderblom, A.Takade, M.Westermark, J.Oscarsson, et al. Vesiclemediated export and assembly of pore-forming oligomers of the enterobacterial ClyA cytotoxin, 2003, Cell, 115, 25–35.
(3)F.J del Castillo, F. Moreno. and I.del Castillo. Secretion of the Escherichia coli K-12 SheA hemolysin is
independent of its cytolytic activity, 2001, FEMS Microbiol.Lett. 204, 281–285.
(4)J.E.Galen, L.Zhao, M.Chinchilla, J.Y.Wang,M.F.Pasetti, J.Green and M.M. Levine. Adaptation of the endogenous Salmonella enterica serovar Typhi clyA-encoded hemolysin for antigen export enhances the immunogenicity of anthrax protective antigen domain 4 expressed by the attenuated live-vector vaccine strain CVD 908-htrA, 2004, Infect. Immun. 72, 7096–7106.
