Anthrax edema factor, voltage-dependent binding to the protective antigen ion channel and comparison to LF binding.
Anthrax toxin complex consists of three different molecules, the binding component protective antigen (PA, 83 kDa), and the enzymatic components lethal factor (LF, 90 kDa) and edema factor (EF, 89 kDa). The 63-kDa N-terminal part of PA, PA(63), forms a heptameric channel that inserts at low pH in endosomal membranes and that is necessary to translocate EF and LF in the cytosol of the target cells. EF is an intracellular active enzyme, which is a calmodulin-dependent adenylate cyclase (89 kDa) that causes a dramatic increase of intracellular cAMP level. Here, the binding of full-length EF on heptameric PA(63) channels was studied in experiments with artificial lipid bilayer membranes. Full-length EF blocks the PA(63) channels in a dose, temperature, voltage, and ionic strength-dependent way with half-saturation constants in the nanomolar concentration range. EF only blocked the PA(63) channels when PA(63) and EF were added to the same side of the membrane, the cis side. Decreasing ionic strength and increasing transmembrane voltage at the cis side of the membranes resulted in a strong decrease of the half-saturation constant for EF binding. This result suggests that ion-ion interactions are involved in EF binding to the PA heptamer. Increasing temperature resulted in increasing half-saturation constants for EF binding to the PA(63) channels. The binding characteristics of EF to the PA(63) channels are compared with those of LF binding. The comparison exhibits similarities but also remarkable differences between the bindings of both toxins to the PA(63) channel.
Neumeyer
Tobias
T
Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum) der Universität Würzburg, Am Hubland, D-97074 Würzburg, Germany.
Tonello
Fiorella
F
Dal Molin
Federica
F
Schiffler
Bettina
B
Benz
Roland
R
D000262:Adenylyl Cyclases / Q000378:metabolism* / Q000494:pharmacology*; D000881:Anthrax / Q000378:metabolism*; D000942:Antigens, Bacterial / Q000378:metabolism* / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000378:metabolism* / Q000494:pharmacology*; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004553:Electric Conductivity; D006863:Hydrogen-Ion Concentration; D007473:Ion Channels / Q000378:metabolism*; D008051:Lipid Bilayers; D009994:Osmolar Concentration; D013696:Temperature
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D007473:Ion Channels; D008051:Lipid Bilayers; C030325:anthrax toxin; D000262:Adenylyl Cyclases