Architecture and assembly of the Gram-positive cell wall

Authors

  • Morgan Beeby,

    1. California Institute of Technology and Howard Hughes Medical Institute, Pasadena, CA, USA
    Current affiliation:
    1. Department of Life Sciences, Imperial College London, South Kensington Campus, London, UK
    Search for more papers by this author
  • James C. Gumbart,

    1. Biosciences Division, Argonne National Laboratory, Argonne, IL, USA
    Current affiliation:
    1. School of Physics, Georgia Institute of Technology, Atlanta, GA, USA
    Search for more papers by this author
  • Benoît Roux,

    1. Biosciences Division, Argonne National Laboratory, Argonne, IL, USA
    2. Department of Biochemistry and Molecular Biology and Gordon Center for Integrative Science, The University of Chicago, Chicago, IL, USA
    Search for more papers by this author
  • Grant J. Jensen

    Corresponding author
    1. California Institute of Technology and Howard Hughes Medical Institute, Pasadena, CA, USA
    • Department of Life Sciences, Imperial College London, South Kensington Campus, London, UK
    Search for more papers by this author

  • Concise description: The architecture of Gram-positive peptidoglycan is still not established, despite decades of research, partially due to a lack of methods to directly visualize it because of it's intermediate size. Here we use electron cryo-tomography to image Gram-positive peptidoglycan in 3D, and combine this with molecular dynamics simulations to bridge scales. Multiple lines of evidence from our work, together with multiple lines of evidence by previous researchers, strongly support the model in which glycan strands run circumferentially around the long axis of the cell.

For correspondence. E-mail jensen@caltech.edu; Tel. (626) 395 8827; Fax (626) 395 5730.

Summary

The bacterial cell wall is a mesh polymer of peptidoglycan – linear glycan strands cross-linked by flexible peptides – that determines cell shape and provides physical protection. While the glycan strands in thin ‘Gram-negative’ peptidoglycan are known to run circumferentially around the cell, the architecture of the thicker ‘Gram-positive’ form remains unclear. Using electron cryotomography, here we show that Bacillus subtilis peptidoglycan is a uniformly dense layer with a textured surface. We further show it rips circumferentially, curls and thickens at free edges, and extends longitudinally when denatured. Molecular dynamics simulations show that only atomic models based on the circumferential topology recapitulate the observed curling and thickening, in support of an ‘inside-to-outside’ assembly process. We conclude that instead of being perpendicular to the cell surface or wrapped in coiled cables (two alternative models), the glycan strands in Gram-positive cell walls run circumferentially around the cell just as they do in Gram-negative cells. Together with providing insights into the architecture of the ultimate determinant of cell shape, this study is important because Gram-positive peptidoglycan is an antibiotic target crucial to the viability of several important rod-shaped pathogens including Bacillus anthracis, Listeria monocytogenes, and Clostridium difficile.

Ancillary