Computational design of transmembrane pores

Nature. 2020 Sep;585(7823):129-134. doi: 10.1038/s41586-020-2646-5. Epub 2020 Aug 26.


Transmembrane channels and pores have key roles in fundamental biological processes1 and in biotechnological applications such as DNA nanopore sequencing2-4, resulting in considerable interest in the design of pore-containing proteins. Synthetic amphiphilic peptides have been found to form ion channels5,6, and there have been recent advances in de novo membrane protein design7,8 and in redesigning naturally occurring channel-containing proteins9,10. However, the de novo design of stable, well-defined transmembrane protein pores that are capable of conducting ions selectively or are large enough to enable the passage of small-molecule fluorophores remains an outstanding challenge11,12. Here we report the computational design of protein pores formed by two concentric rings of α-helices that are stable and monodisperse in both their water-soluble and their transmembrane forms. Crystal structures of the water-soluble forms of a 12-helical pore and a 16-helical pore closely match the computational design models. Patch-clamp electrophysiology experiments show that, when expressed in insect cells, the transmembrane form of the 12-helix pore enables the passage of ions across the membrane with high selectivity for potassium over sodium; ion passage is blocked by specific chemical modification at the pore entrance. When incorporated into liposomes using in vitro protein synthesis, the transmembrane form of the 16-helix pore-but not the 12-helix pore-enables the passage of biotinylated Alexa Fluor 488. A cryo-electron microscopy structure of the 16-helix transmembrane pore closely matches the design model. The ability to produce structurally and functionally well-defined transmembrane pores opens the door to the creation of designer channels and pores for a wide variety of applications.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Cell Line
  • Computer Simulation*
  • Cryoelectron Microscopy
  • Crystallography, X-Ray
  • Electric Conductivity
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Genes, Synthetic / genetics*
  • Hydrazines
  • Ion Channels / chemistry*
  • Ion Channels / genetics*
  • Ion Channels / metabolism
  • Ion Transport
  • Liposomes / metabolism
  • Models, Molecular*
  • Patch-Clamp Techniques
  • Porins / chemistry
  • Porins / genetics
  • Porins / metabolism
  • Protein Engineering
  • Protein Structure, Secondary
  • Solubility
  • Synthetic Biology*
  • Water / chemistry


  • Alexa 488 hydrazide
  • Hydrazines
  • Ion Channels
  • Liposomes
  • Porins
  • Water