Listeriolysin O, Recombinant (LLO)

Listeriolysin O (aka LLO) is a hemolysin produced by Listeria monocytogenes bacteria, the pathogen responsible for causing listeriosis. The toxin may be regarded as a virulence factor, since it is crucial for the virulence of L. monocytogenes. LLO is a single polypeptide protein encoded by the hlyA gene and composed of 529 residues. LLO is a thiol-activated cholesterol-dependent pore forming toxin protein; therefore, it is activated by reducing agents and inhibited by oxidizing agents. Still, LLO differs from other thiol-activated toxins, as its cytolytic activity is maximized at a pH of 5.5. Inside the acidic phagosomes (average pH ~ 5.9) of cells that have phagocytosed L. monocytogenes, LLO is selectively activated by maximizing activity at a pH of 5.5. Following the phagosome lysis by LLO, the bacterium breaks out into the cytosol, where it is able to grow intracellularly, and the toxin has reduced activity in the more basic cytosol. Thus, LLO permits L. monocytogenes to break out from the phagosomes into the cytosol without harming the plasma membrane of the infected cell, which allows the bacteria to live intracellularly, where they are sheltered from extracellular immune system factors such as the complement system and antibodies. LLO also brings about dephosphorylation of histone H3 and deacetylation of histone H4 in the early phases of infection, before entry of L. monocytogenes into the host cell. The pore-forming activity is not implicated in causing the histone modifications. The modifications of the histones affect the down regulation of genes encoding proteins involved in the inflammatory response. Therefore, LLO may be significant in subverting the host immune response to L. monocytogenes. At its NH2-terminus it possesses a 25 residues long typical signal sequence excited during the secretion process. Moreover, in its NH2-terminus there is also a 19 amino acids PEST- like sequence that may target this toxin for degradation. The PEST-like sequence found in LLO and is considered crucial for virulence, given that mutants lacking the sequence lysed the host cell. Nevertheless, contrary to PEST's supposed role in protein degradation, evidence implies that the PEST-like sequence may control LLO production in the cytosol rather than increase degradation of LLO.\n\nBiological Activity:\nRecombinant LLO has a hemolytic activity 1.25x10e6 HU/mg. 2mM DTT is used to reactivate the toxin.\n\nApplications:\nSuitable for use in Cytosolic delivery of molecules, peptides, oligonucleodides and plasmid DNA, Production of specific monoclonal antibodies and Detection of anti-listeriolysin O antibodies. Other applications not tested.\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20 degrees C. Aliquots are stable for 6 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.

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SPECIFICATIONS

Catalog Number

L2650-50

Size

10ug

Form

Supplied as a liquid in 50mM NaH2PO4, 0.5M sodium chloride, 1mM EDTA, 2.7mM potassium chloride, 1mM DTT, 5% glycerol.

Purity

(same/more than)90% by SDS-PAGE.

References

1. Rocort, J., anddH2O Cossart, P. (1997) (Doyle, M.P., and Montville, T.J., Ed.), pp.337-352, ASM press, Washington D.C. 2. Palmer, M. (2001)Toxicon 39, 1681-1689. 3. Giammarini C., Andreoni F., Amagliani G., Casiere A., Barocci S. and Magnani M. (2003) Protein Expr. Purif. 28: 78-85. 4. Provoda C.J. and Lee K.-D. (2000) Adv. Drug. Deliv. Rev. 41: 209-221. 5. Nato F., Reich K., Lhopital S., Rouyre S., Geoffrey C., Mazie J.C. and Cossart P. (1991) Infect Immun. 59:4641-4646. 6. Barbuddhe S. B., Malik S. V. S. and Gupta L.K. (2000) J. Veterinary Medicine. 47: 497- 502.