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A kinetic analysis of protein transport through the anthrax toxin channel.

Basilio D, Kienker PK, Briggs SW, Finkelstein A - J. Gen. Physiol. (2011)

Bottom Line: 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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology and Biophysics, Albert Einstein College of Medicine, Bronx, NY 10461, USA. dab2043@-med.cornell.edu

ABSTRACT
Anthrax toxin is composed of three proteins: a translocase heptameric channel, (PA(63))(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. (PA(63))(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(N)s. 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.

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The dimensionless half-time for translocation (tD/L2) as a function of the Péclet number, Ω, calculated from the drift-diffusion model.
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fig5: The dimensionless half-time for translocation (tD/L2) as a function of the Péclet number, Ω, calculated from the drift-diffusion model.

Mentions: Each model W(Ω, t) curve can be described by two parameters, u and Ω. For a given mobility, the rise of W(Ω, t) with time becomes faster, and thus the half-times become shorter, with increasing Ω (Fig. 5). We sometimes found it more convenient to express W(Ω, t) in terms of t1/2 (which is proportional to 1/u) and Ω. The half-time can be determined directly from the data curve (normalized conductance vs. time). To facilitate the fitting of Ω, we used a rescaled time variable for each curve, t′ = t/t1/2, to make the model and data curves superimpose at a value of 1/2 when t′ = 1. We then plotted a family of model curves for various values of Ω and visually determined which one best matched a given data curve. For this, we emphasized the earlier part of the curve on the rationale that the later part was more likely to be contaminated by channel-gating effects. All analysis routines were written using the program Igor Pro (WaveMetrics, Inc.).


A kinetic analysis of protein transport through the anthrax toxin channel.

Basilio D, Kienker PK, Briggs SW, Finkelstein A - J. Gen. Physiol. (2011)

The dimensionless half-time for translocation (tD/L2) as a function of the Péclet number, Ω, calculated from the drift-diffusion model.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3105512&req=5

fig5: The dimensionless half-time for translocation (tD/L2) as a function of the Péclet number, Ω, calculated from the drift-diffusion model.
Mentions: Each model W(Ω, t) curve can be described by two parameters, u and Ω. For a given mobility, the rise of W(Ω, t) with time becomes faster, and thus the half-times become shorter, with increasing Ω (Fig. 5). We sometimes found it more convenient to express W(Ω, t) in terms of t1/2 (which is proportional to 1/u) and Ω. The half-time can be determined directly from the data curve (normalized conductance vs. time). To facilitate the fitting of Ω, we used a rescaled time variable for each curve, t′ = t/t1/2, to make the model and data curves superimpose at a value of 1/2 when t′ = 1. We then plotted a family of model curves for various values of Ω and visually determined which one best matched a given data curve. For this, we emphasized the earlier part of the curve on the rationale that the later part was more likely to be contaminated by channel-gating effects. All analysis routines were written using the program Igor Pro (WaveMetrics, Inc.).

Bottom Line: 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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology and Biophysics, Albert Einstein College of Medicine, Bronx, NY 10461, USA. dab2043@-med.cornell.edu

ABSTRACT
Anthrax toxin is composed of three proteins: a translocase heptameric channel, (PA(63))(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. (PA(63))(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(N)s. 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.

Show MeSH
Related in: MedlinePlus