Peptide Permeation across a Phosphocholine Membrane: An Atomically Detailed Mechanism Determined through Simulations and Supported by Experimentation

Alfredo E. Cardenas; Chad I. Drexler; Rachel Nechushtai; Ron Mittler; Assaf Friedler; Lauren J. Webb; Ron Elber

doi:10.1021/acs.jpcb.1c10966

Peptide Permeation across a Phosphocholine Membrane: An Atomically Detailed Mechanism Determined through Simulations and Supported by Experimentation

Alfredo E. Cardenas, Chad I. Drexler, Rachel Nechushtai, Ron Mittler, Assaf Friedler, Lauren J. Webb^*, Ron Elber^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

11 Scopus citations

Abstract

Cell-penetrating peptides (CPPs) facilitate translocation across biological membranes and are of significant biological and medical interest. Several CPPs can permeate into specific cells and organelles. We examine the incorporation and translocation of a novel anticancer CPP in a dioleoylphosphatidylcholine (DOPC) lipid bilayer membrane. The peptide, NAF-144-67, is a short fragment of a transmembrane protein, consisting of hydrophobic N-terminal and charged C-terminal segments. Experiments using fluorescently labeled NAF-144-67 in ∼100 nm DOPC vesicles and atomically detailed simulations conducted with Milestoning support a model in which a significant barrier for peptide-membrane entry is found at the interface between the aqueous solution and membrane. The initial step is the insertion of the N-terminal segment and the hydrophobic helix into the membrane, passing the hydrophilic head groups. Both experiments and simulations suggest that the free energy difference in the first step of the permeation mechanism in which the hydrophobic helix crosses the phospholipid head groups is -0.4 kcal mol-1 slightly favoring motion into the membrane. Milestoning calculations of the mean first passage time and the committor function underscore the existence of an early polar barrier followed by a diffusive barrierless motion in the lipid tail region. Permeation events are coupled to membrane fluctuations that are examined in detail. Our study opens the way to investigate in atomistic resolution the molecular mechanism, kinetics, and thermodynamics of CPP permeation to diverse membranes.

Original language	American English
Pages (from-to)	2834-2849
Number of pages	16
Journal	Journal of Physical Chemistry B
Volume	126
Issue number	15
DOIs	https://doi.org/10.1021/acs.jpcb.1c10966
State	Published - 21 Apr 2022

Bibliographical note

Funding Information:
This research was supported by NIH, Grant No. GM 59796 (to R.E.) and GM 111364 (to R.E., L.J.W., R.M.); the Welch Foundation, Grant No. F-1896 (to R.E.) and F-1722 (to L.J.W.); and the BSF grant number 2020094 to R.N., A.F., L.J.W., and R.E. Part of the computations were done using HPC resources from the Texas Advanced Computing Center (TACC) at the University of Texas at Austin. The authors gratefully acknowledge the use of facilities at the Texas Materials Institute at the University of Texas at Austin.

Publisher Copyright:
© 2022 American Chemical Society.

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title = "Peptide Permeation across a Phosphocholine Membrane: An Atomically Detailed Mechanism Determined through Simulations and Supported by Experimentation",

abstract = "Cell-penetrating peptides (CPPs) facilitate translocation across biological membranes and are of significant biological and medical interest. Several CPPs can permeate into specific cells and organelles. We examine the incorporation and translocation of a novel anticancer CPP in a dioleoylphosphatidylcholine (DOPC) lipid bilayer membrane. The peptide, NAF-144-67, is a short fragment of a transmembrane protein, consisting of hydrophobic N-terminal and charged C-terminal segments. Experiments using fluorescently labeled NAF-144-67 in ∼100 nm DOPC vesicles and atomically detailed simulations conducted with Milestoning support a model in which a significant barrier for peptide-membrane entry is found at the interface between the aqueous solution and membrane. The initial step is the insertion of the N-terminal segment and the hydrophobic helix into the membrane, passing the hydrophilic head groups. Both experiments and simulations suggest that the free energy difference in the first step of the permeation mechanism in which the hydrophobic helix crosses the phospholipid head groups is -0.4 kcal mol-1 slightly favoring motion into the membrane. Milestoning calculations of the mean first passage time and the committor function underscore the existence of an early polar barrier followed by a diffusive barrierless motion in the lipid tail region. Permeation events are coupled to membrane fluctuations that are examined in detail. Our study opens the way to investigate in atomistic resolution the molecular mechanism, kinetics, and thermodynamics of CPP permeation to diverse membranes.",

author = "Cardenas, {Alfredo E.} and Drexler, {Chad I.} and Rachel Nechushtai and Ron Mittler and Assaf Friedler and Webb, {Lauren J.} and Ron Elber",

note = "Funding Information: This research was supported by NIH, Grant No. GM 59796 (to R.E.) and GM 111364 (to R.E., L.J.W., R.M.); the Welch Foundation, Grant No. F-1896 (to R.E.) and F-1722 (to L.J.W.); and the BSF grant number 2020094 to R.N., A.F., L.J.W., and R.E. Part of the computations were done using HPC resources from the Texas Advanced Computing Center (TACC) at the University of Texas at Austin. The authors gratefully acknowledge the use of facilities at the Texas Materials Institute at the University of Texas at Austin. Publisher Copyright: {\textcopyright} 2022 American Chemical Society.",

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AU - Friedler, Assaf

AU - Webb, Lauren J.

AU - Elber, Ron

N1 - Funding Information: This research was supported by NIH, Grant No. GM 59796 (to R.E.) and GM 111364 (to R.E., L.J.W., R.M.); the Welch Foundation, Grant No. F-1896 (to R.E.) and F-1722 (to L.J.W.); and the BSF grant number 2020094 to R.N., A.F., L.J.W., and R.E. Part of the computations were done using HPC resources from the Texas Advanced Computing Center (TACC) at the University of Texas at Austin. The authors gratefully acknowledge the use of facilities at the Texas Materials Institute at the University of Texas at Austin. Publisher Copyright: © 2022 American Chemical Society.

PY - 2022/4/21

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N2 - Cell-penetrating peptides (CPPs) facilitate translocation across biological membranes and are of significant biological and medical interest. Several CPPs can permeate into specific cells and organelles. We examine the incorporation and translocation of a novel anticancer CPP in a dioleoylphosphatidylcholine (DOPC) lipid bilayer membrane. The peptide, NAF-144-67, is a short fragment of a transmembrane protein, consisting of hydrophobic N-terminal and charged C-terminal segments. Experiments using fluorescently labeled NAF-144-67 in ∼100 nm DOPC vesicles and atomically detailed simulations conducted with Milestoning support a model in which a significant barrier for peptide-membrane entry is found at the interface between the aqueous solution and membrane. The initial step is the insertion of the N-terminal segment and the hydrophobic helix into the membrane, passing the hydrophilic head groups. Both experiments and simulations suggest that the free energy difference in the first step of the permeation mechanism in which the hydrophobic helix crosses the phospholipid head groups is -0.4 kcal mol-1 slightly favoring motion into the membrane. Milestoning calculations of the mean first passage time and the committor function underscore the existence of an early polar barrier followed by a diffusive barrierless motion in the lipid tail region. Permeation events are coupled to membrane fluctuations that are examined in detail. Our study opens the way to investigate in atomistic resolution the molecular mechanism, kinetics, and thermodynamics of CPP permeation to diverse membranes.

AB - Cell-penetrating peptides (CPPs) facilitate translocation across biological membranes and are of significant biological and medical interest. Several CPPs can permeate into specific cells and organelles. We examine the incorporation and translocation of a novel anticancer CPP in a dioleoylphosphatidylcholine (DOPC) lipid bilayer membrane. The peptide, NAF-144-67, is a short fragment of a transmembrane protein, consisting of hydrophobic N-terminal and charged C-terminal segments. Experiments using fluorescently labeled NAF-144-67 in ∼100 nm DOPC vesicles and atomically detailed simulations conducted with Milestoning support a model in which a significant barrier for peptide-membrane entry is found at the interface between the aqueous solution and membrane. The initial step is the insertion of the N-terminal segment and the hydrophobic helix into the membrane, passing the hydrophilic head groups. Both experiments and simulations suggest that the free energy difference in the first step of the permeation mechanism in which the hydrophobic helix crosses the phospholipid head groups is -0.4 kcal mol-1 slightly favoring motion into the membrane. Milestoning calculations of the mean first passage time and the committor function underscore the existence of an early polar barrier followed by a diffusive barrierless motion in the lipid tail region. Permeation events are coupled to membrane fluctuations that are examined in detail. Our study opens the way to investigate in atomistic resolution the molecular mechanism, kinetics, and thermodynamics of CPP permeation to diverse membranes.

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ER -

Peptide Permeation across a Phosphocholine Membrane: An Atomically Detailed Mechanism Determined through Simulations and Supported by Experimentation

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