title: Small-angle X-ray scattering for imaging of surface layers on intact bacteria in the native environment.
abstract: Crystalline cell surface layers (S-layers) represent a natural two-dimensional (2D) protein self-assembly system with nanometer-scale periodicity that decorate many prokaryotic cells. Here, we analyze the S-layer on intact bacterial cells of the Gram-positive organism Geobacillus stearothermophilus ATCC 12980 and the Gram-negative organism Aquaspirillum serpens MW5 by small-angle X-ray scattering (SAXS) and relate it to the structure obtained by transmission electron microscopy (TEM) after platinum/carbon shadowing. By measuring the scattering pattern of X rays obtained from a suspension of bacterial cells, integral information on structural elements such as the thickness and lattice parameters of the S-layers on intact, hydrated cells can be obtained nondestructively. In contrast, TEM of whole mounts is used to analyze the S-layer lattice type and parameters as well as the physical structure in a nonaqueous environment and local information on the structure is delivered. Application of SAXS to S-layer research on intact bacteria is a challenging task, as the scattering volume of the generally thin (3- to 30-nm) bacterial S-layers is low in comparison to the scattering volume of the bacterium itself. For enhancement of the scattering contrast of the S-layer in SAXS measurement, either silicification (treatment with tetraethyl orthosilicate) is used, or the difference between SAXS signals from an S-layer-deficient mutant and the corresponding S-layer-carrying bacterium is used for determination of the scattering signal. The good agreement of the SAXS and TEM data shows that S-layers on the bacterial cell surface are remarkably stable.
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Vienna, Austria.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Sekot
authors.forename: David
authors.initials: D
authors.lastname: Schuster
authors.forename: Paul
authors.initials: P
authors.lastname: Messner
authors.forename: Dietmar
authors.initials: D
authors.lastname: Pum
authors.forename: Herwig
authors.initials: H
authors.lastname: Peterlik
authors.forename: Christina
authors.initials: C
authors.lastname: Schäffer
mesh_terms: D001419:Bacteria / Q000648:ultrastructure; D042621:Comamonadaceae; D001411:Geobacillus stearothermophilus; D046529:Microscopy, Electron, Transmission; D053838:Scattering, Small Angle; D014965:X-Rays