Anthrax toxin-mediated delivery of cholera toxin-A subunit into the cytosol of mammalian cells.
The protective antigen (PA) component of anthrax toxin mediates delivery of either lethal factor (LF) or oedema factor into the cytosol of mammalian cells. The N-terminal domain of LF(1-254) (amino acids 1-254 of LF) binds to PA and, when fused to heterologous proteins, delivers such proteins into the cytosol. In the present study, we fused the catalytic subunit of cholera toxin (CT-A) with LF(1-254) and showed that the fusion protein LF(1-254)-CT-A retains ADP-ribosylation activity in solution and increased intracellular cAMP levels in J774A.1 macrophage cells when added together with PA. A mutant fusion protein, in which arginine-7 of CT-A was replaced with lysine, did not show ADP-ribosylation activity in solution and failed to increase cAMP levels in macrophage cells. The data show that LF(1-254)-CT-A retains its catalytic activity in solution as well as when translocated into the cytosol of eukaryotic cells via an alternative pathway to the GM(1) receptor used by CT.
Centre for Biochemical Technology, Mall Road, Near Jubilee Hall, Delhi-1 10007, India.
M
M
Sharma
H
H
Khanna
N
N
Arora
Y
Y
Singh
D000244:Adenosine Diphosphate / Q000378:metabolism; D000818:Animals; D000942:Antigens, Bacterial; D001427:Bacterial Toxins / Q000008:administration & dosage / Q000235:genetics*; D002772:Cholera Toxin / Q000008:administration & dosage / Q000235:genetics*; D000242:Cyclic AMP / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects* / Q000378:metabolism; D004337:Drug Carriers; D016503:Drug Delivery Systems; D008322:Mammals; D009154:Mutation; D011956:Receptors, Cell Surface / Q000187:drug effects
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D004337:Drug Carriers; D011956:Receptors, Cell Surface; C030325:anthrax toxin; D000244:Adenosine Diphosphate; D002772:Cholera Toxin; D000242:Cyclic AMP