A kinetic analysis of protein transport through the anthraxtoxin channel

Anthrax toxin is composed of three proteins: a translocase heptameric channel, (PA₆₃)7, formed from protective antigen (PA), which allows the other two proteins, lethal factor (LF) and edema factor (EF), to translocate across a host cell's endosomal membrane, disrupting cellular homeostasis. (PA₆₃)₇ incorporated into planar phospholipid bilayer membranes forms a channel capable of transporting LF and EF. Protein translocation through the channel can be driven by voltage on a timescale of seconds. A characteristic of the translocation of LF_N, the N-terminal 263 residues of LF, is its S-shaped kinetics. Because all of the translocation experiments reported in the literature have been performed with more than one LF_N molecule bound to most of the channels, it is not clear whether the S-shaped kinetics are an intrinsic characteristic of translocation kinetics or are merely a consequence of the translocation in tandem of two or three LF_Ns. In this paper, we show both in macroscopic and single-channel experiments that even with only one LF_N bound to the channel, the translocation kinetics are S shaped. As expected, the translocation rate is slower with more than one LF_N bound. We also present a simple electrodiffusion model of translocation in which LF_N is represented as a charged rod that moves subject to both Brownian motion and an applied electric field. The cumulative distribution of first-passage times of the rod past the end of the channel displays S-shaped kinetics with a voltage dependence in agreement with experimental data.