Photolithographic Fabrication of Microapertures with Well-Defined, Three-Dimensional Geometries for Suspended Lipid Membrane Studies

Abstract The amino acid sequence of a protein encodes information on its three-dimensional structure and specific functionality. De novo protein design has emerged as a method to manipulate the primary structure for the development of artificial proteins and peptides with desired functionality. This paper describes the de novo design of a pore-forming peptide that has a β-hairpin structure and assembles to form a stable nanopore in a bilayer lipid membrane. This large synthetic nanopore is an entirely artificial device with practical applications. This peptide, named SV28, forms nanopore structures ranging from 1.6 to 6.2 nm in diameter assembled from 7 to 18 monomers. The nanopore formed with a diameter of 5 nm is able to detect long double-stranded DNA (dsDNA) with 1 kbp length. Moreover, the larger sized nanopore can discriminate and human telomeric DNA (G-quadruplex, G4). The blocking current signals allowed us to investigate the translocation behavior of dsDNA or G4 structure at the single molecule level. Such de novo design of peptide sequences has the potential to create novel nanopores, which would be applicable in molecular transporter between across lipid membrane.

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De Novo Design of a Nanopore for DNA Detection Incorporating a β-hairpin Peptide

10.26434/chemrxiv.12286337.v1 ◽

2020 ◽

Author(s):

Keisuke Shimizu ◽

Batsaikhan Mijiddorj ◽

Shuhei Yoshida ◽

Shiori Akayama ◽

Yoshio Hamada ◽

...

Keyword(s):

Single Molecule ◽

Protein Design ◽

Lipid Membrane ◽

De Novo ◽

Three Dimensional ◽

Bilayer Lipid Membrane ◽

De Novo Design ◽

Dimensional Structure ◽

Practical Applications ◽

Single Molecule Level

The amino acid sequence of a protein encodes information on its three-dimensional structure and specific functionality. De novo protein design has emerged as a method to manipulate the primary structure for the development of artificial proteins and peptides with desired functionality. This paper describes the de novo design of a pore-forming peptide that has a β-hairpin structure and assembles to form a stable nanopore in a bilayer lipid membrane. This large synthetic nanopore is an entirely artificial device with practical applications. This peptide, named SV28, forms nanopore structures ranging from 1.6 to 6.2 nm in diameter assembled from 7 to 18 monomers. The nanopore formed with a diameter of 5 nm is able to detect long double-stranded DNA (dsDNA) with 1 kbp length, and measurement of current signals allowed us to investigate the translocation behavior of dsDNA at the single molecule level. Such de novo design of peptide sequences has the potential to create assembled structure in lipid membrane such as novel nanopores, which would also be applicable in molecular transporter between inside and outside of lipid membrane.

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The insertion of human apolipoprotein H into phospholipid membranes: a monolayer study

Biochemical Journal ◽

10.1042/bj3350225 ◽

1998 ◽

Vol 335 (2) ◽

pp. 225-232 ◽

Cited By ~ 35

Author(s):

Shao-Xiong WANG ◽

Guo-ping CAI ◽

Sen-fang SUI

Keyword(s):

Lipid Membrane ◽

Hydrophobic Interactions ◽

Three Dimensional ◽

Dimensional Structure ◽

Heat Inactivation ◽

Sequence Motif ◽

Phospholipid Membranes ◽

Human Apolipoprotein ◽

Stable Domain ◽

Apolipoprotein H

Apolipoprotein H (ApoH) is a plasma glycoprotein isolated from human serum. The interactions of ApoH with lipid membrane were reported to be essential for its physiological and pathogenic roles. In this paper we studied the ability of ApoH to insert into phospholipid membranes using the monolayer approach. The results show that ApoH is surface active and can insert into the lipid monolayers. The insertion ability of ApoH is stronger when a higher content of negatively charged lipids is present in the membrane. The acidic-pH and low-ionic-strength conditions will also enhance ApoH insertion, but these factors may not have much influence on the final insertion ability of ApoH, suggesting that, in the mechanism of ApoH insertion, not only electrostatic forces, but also hydrophobic interactions, are evidently involved. Modification by heat inactivation and reduction/alkylation does not change the critical insertion pressure (πc) of ApoH, suggesting a stable domain, maybe a linear sequence motif, but not the native three-dimensional structure of ApoH, is responsible for its insertion. The extent to which insertion of ApoH into phospholipid membranes may facilitate the ‘immune cleaning ’ of plasma liposomes is discussed.

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De novo design of a nanopore for single-molecule detection that incorporates a β-hairpin peptide

Nature Nanotechnology ◽

10.1038/s41565-021-01008-w ◽

2021 ◽

Author(s):

Keisuke Shimizu ◽

Batsaikhan Mijiddorj ◽

Masataka Usami ◽

Ikuro Mizoguchi ◽

Shuhei Yoshida ◽

...

Keyword(s):

Single Molecule ◽

Lipid Membrane ◽

De Novo ◽

Three Dimensional ◽

Bilayer Lipid Membrane ◽

De Novo Design ◽

Dimensional Structure ◽

Practical Applications ◽

Single Polypeptide Chain ◽

Single Polypeptide

AbstractThe amino-acid sequence of a protein encodes information on its three-dimensional structure and specific functionality. De novo design has emerged as a method to manipulate the primary structure for the development of artificial proteins and peptides with desired functionality. This paper describes the de novo design of a pore-forming peptide, named SV28, that has a β-hairpin structure and assembles to form a stable nanopore in a bilayer lipid membrane. This large synthetic nanopore is an entirely artificial device for practical applications. The peptide forms multidispersely sized nanopore structures ranging from 1.7 to 6.3 nm in diameter and can detect DNAs. To form a monodispersely sized nanopore, we redesigned the SV28 by introducing a glycine-kink mutation. The resulting redesigned peptide forms a monodisperse pore with a diameter of 1.7 nm leading to detection of a single polypeptide chain. Such de novo design of a β-hairpin peptide has the potential to create artificial nanopores, which can be size adjusted to a target molecule.

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De Novo Design of a Nanopore for DNA Detection Incorporating a β-hairpin Peptide

10.26434/chemrxiv.12286337 ◽

2020 ◽

Author(s):

Keisuke Shimizu ◽

Batsaikhan Mijiddorj ◽

Shuhei Yoshida ◽

Shiori Akayama ◽

Yoshio Hamada ◽

...

Keyword(s):

Single Molecule ◽

Protein Design ◽

Lipid Membrane ◽

De Novo ◽

Three Dimensional ◽

Bilayer Lipid Membrane ◽

De Novo Design ◽

Dimensional Structure ◽

Practical Applications ◽

Single Molecule Level

The amino acid sequence of a protein encodes information on its three-dimensional structure and specific functionality. De novo protein design has emerged as a method to manipulate the primary structure for the development of artificial proteins and peptides with desired functionality. This paper describes the de novo design of a pore-forming peptide that has a β-hairpin structure and assembles to form a stable nanopore in a bilayer lipid membrane. This large synthetic nanopore is an entirely artificial device with practical applications. This peptide, named SV28, forms nanopore structures ranging from 1.6 to 6.2 nm in diameter assembled from 7 to 18 monomers. The nanopore formed with a diameter of 5 nm is able to detect long double-stranded DNA (dsDNA) with 1 kbp length, and measurement of current signals allowed us to investigate the translocation behavior of dsDNA at the single molecule level. Such de novo design of peptide sequences has the potential to create assembled structure in lipid membrane such as novel nanopores, which would also be applicable in molecular transporter between inside and outside of lipid membrane.

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De Novo Design of a Nanopore for DNA Detection Incorporating a β-hairpin Peptide

10.26434/chemrxiv.12286337.v2 ◽

2020 ◽

Cited By ~ 1

Author(s):

Keisuke Shimizu ◽

Batsaikhan Mijiddorj ◽

Shuhei Yoshida ◽

Shiori Akayama ◽

Yoshio Hamada ◽

...

Keyword(s):

Single Molecule ◽

Protein Design ◽

Lipid Membrane ◽

De Novo ◽

Three Dimensional ◽

Bilayer Lipid Membrane ◽

De Novo Design ◽

Dimensional Structure ◽

Practical Applications ◽

Single Molecule Level

The amino acid sequence of a protein encodes information on its three-dimensional structure and specific functionality. De novo protein design has emerged as a method to manipulate the primary structure for the development of artificial proteins and peptides with desired functionality. This paper describes the de novo design of a pore-forming peptide that has a β-hairpin structure and assembles to form a stable nanopore in a bilayer lipid membrane. This large synthetic nanopore is an entirely artificial device with practical applications. This peptide, named SV28, forms nanopore structures ranging from 1.6 to 6.2 nm in diameter assembled from 7 to 18 monomers. The nanopore formed with a diameter of 5 nm is able to detect long double-stranded DNA (dsDNA) with 1 kbp length, and measurement of current signals allowed us to investigate the translocation behavior of dsDNA at the single molecule level. Such de novo design of peptide sequences has the potential to create assembled structure in lipid membrane such as novel nanopores, which would also be applicable in molecular transporter between inside and outside of lipid membrane.

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Settling of a vesicle in the limit of quasispherical shapes

Journal of Fluid Mechanics ◽

10.1017/jfm.2011.427 ◽

2011 ◽

Vol 690 ◽

pp. 227-261 ◽

Cited By ~ 19

Author(s):

G. Boedec ◽

M. Jaeger ◽

M. Leonetti

Keyword(s):

Lipid Membrane ◽

Initial Conditions ◽

Three Dimensional ◽

Small Deformation ◽

Thermal Fluctuations ◽

Bond Number ◽

Membrane Area ◽

Final Shape ◽

Curvature Energy ◽

Internal Volume

AbstractVesicles are drops of radius of a few tens of micrometres bounded by an impermeable lipid membrane of approximately 4 nm thickness in a viscous fluid. The salient characteristics of such a deformable object are a membrane rigidity governed by flexion due to curvature energy and a two-dimensional membrane fluidity characterized by a local membrane incompressibility. This provides unique properties with strong constraints on the internal volume and membrane area. Yet, when subjected to external stresses, vesicles exhibit a large deformability. The deformation of a settling vesicle in an infinite flow is studied theoretically, assuming a quasispherical shape and expanding all variables of the problem onto spherical harmonics. The contribution of thermal fluctuations is neglected in this analysis. A system of equations describing the temporal evolution of the shape is derived with this formalism. The final shape and the settling velocity are then determined and depend on two dimensionless parameters: the Bond number and the excess area. This simultaneous study leads to three stationary shapes, an egg-like shape already observed in an analogous experimental configuration in the limit of weak flow magnitude (Chatkaew, Georgelin, Jaeger & Leonetti, Phys. Rev. Lett, 2009, vol. 103(24), 248103), a parachute-like shape and a non-trivial non-axisymmetrical shape. The final shape depends on the initial conditions: prolate or oblate vesicle and orientation compared with gravity. The analytical solution in the small deformation regime is compared with numerical results obtained with a three-dimensional code. A very good agreement between numerical and theoretical results is found.

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