Monolayer freeze‐fracture and scanning tunneling microscopy

Fisher, Knute A.

doi:10.1002/jemt.1060130408

Cited by 12 publications

(1 citation statement)

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“…Since it is not yet possible to distinguish these potential artifacts from specimen structure, considerable care is needed to interpret topographical detail of biological surfaces. An important control for assessing the quality of topographical detail is to utilize structural landmarks or barrier heights for the positive identification of biomacromolecules in scanned images (15,25). Highly ordered biological surfaces (10,16,27,36,37), such as the paracrystalline proteinaceous structures examined in this study, provide mechanically rigid surfaces that contain useful landmark features for assessing image quality.…”

Section: Resultsmentioning

confidence: 99%

Transmission electron microscopy, scanning tunneling microscopy, and atomic force microscopy of the cell envelope layers of the archaeobacterium Methanospirillum hungatei GP1

et al. 1993

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Methanospirllum hungatei GP1 possesses paracrystalline cell envelope components including end plugs and a sheath formed from stacked hoops. Both negative-stain transmission electron microscopy (TEM) and scanning tunneling microscopy (STM) distinguished the 2.8-nm repeat on the outer surface of the sheath, while negative-stain TEM alone demonstrated this repeat around the outer circumference of individual hoops. Thin sections revealed a wave-like outer sheath surface, while STM showed the presence of deep grooves that precisely defined the hoop-to-hoop boundaries at the waveform nodes. Atomic force microscopy of sheath tubes containing entrapped end plugs emphasized the end plug structure, suggesting that the sheath was malleable enough to collapse over the end plugs and deform to mimic the shape of the underlying structure. High-resolution atomic force microscopy has revised the former idea of end plug structure so that we believe each plug consists of at least four discs, each of which is -3.5 nm thick. Pt shadow TEM and STM both demonstrated the 14-nm hexagonal, particulate surface of an end plug, and STM showed the constituent particles to be lobed structures with numerous smaller projections, presumably corresponding to the molecular folding of the particle.All current techniques for ultrahigh resolution of biomolecular structure have inherent drawbacks. For example, not only does transmission electron microscopy (TEM) usually require heavy metal contrasting agents, it also produces high energy loads and high vacuums on the specimen. Only in exceptional cases, such as the purple membrane of Halobacterium spp. (18), have molecular folding data been obtained. The inception of scanning tunneling microscopy (STM) (7) and atomic force microscopy (AFM) (6) has certainly made the atomic resolution of hard, inanimate surfaces feasible, and there is a good possibility that this same resolution can be approached in biology. STM and AFM are currently being used on a number of biostructures and their constituent biopolymers, but they are still relatively new techniques in structural biology, and submolecular resolution must be interpreted with caution since biomaterials are loosely bonded and easily deformable. Specimens must be chosen with care, and close attention must be paid to the possibility of induced artifacts. It is best to take a multitechnique approach which combines high-resolution methodologies based on different principles; uniformity of high-resolution detail from each ensures accuracy of interpretation. Using this rationale, we have combined TEM, STM, and AFM to study paracrystalline surfaces possessed by the archaeobacterium Methanospirillum hungatei.STM and AFM rely on the raster scanning of a fine-tip probe over a surface, with piezoelectric ceramics to control movement to within subnanometer distances, which forms a topographical three-dimensional image of the specimen. In STM, the vertical tip displacement during scanning is depen-* Corresponding author. dent on the tunneling current between the pen...

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