Structure of purple membrane from halobacterium halobium: recording, measurement and evaluation of electron micrographs at 3.5 Å resolution

Henderson, Richard A.; Baldwin, Joyce; Downing, Kenneth H.; Lepault, Jean; Zemlin, F.

doi:10.1016/0304-3991(86)90203-2

Cited by 710 publications

(509 citation statements)

References 35 publications

Supporting

Mentioning

499

Contrasting

Unclassified

Order By: Relevance

“…Suitable areas for Fourier filtering and averaging were computationally unbend using the MRC-based SPECTRA platform (Schmid et al, 1993), Fourier filtered, and averaged using the MRC (Henderson et al, 1986) and SUPRIM packages (Schroeter and Bretaudiere, 1996). The Fourier filtered images and averages in Figure 4B and C were obtained assuming lattice parameters with 16 nm axial repeats (corresponding to two consecutive tubulin dimers along the protofilament axis).…”

Section: Image Analysismentioning

confidence: 99%

Surface Topography of Microtubule Walls Decorated with Monomeric and Dimeric Kinesin Constructs

Hoenger

Doerhoefer²,

Woehlke³

et al. 2000

Biological Chemistry

View full text Add to dashboard Cite

The surface topography of opened-up microtubule walls (sheets) decorated with monomeric and dimeric kinesin motor domains was investigated by freezedrying and unidirectional metal shadowing. Electron microscopy of surface-shadowed specimens produces images with a high signal/noise ratio, which enable a direct observation of surface features below 2 nm detail. Here we investigate the inner and outer surface of microtubules and tubulin sheets with and without decoration by kinesin motor domains. Tubulin sheets are flattened walls of microtubules, keeping lateral protofilament contacts intact. Surface shadowing reveals the following features: (i) when the microtubule outside is exposed the surface relief is dominated by the bound motor domains. Monomeric motor constructs generate a strong 8 nm periodicity, corresponding to the binding of one motor domain per ␣-␤-tubulin heterodimer. This surface periodicity largely disappears when dimeric kinesin motor domains are used for decoration, even though it is still visible in negatively stained or frozen hydrated specimens. This could be explained by disorder in the binding of the second (loosely tethered) kinesin head, and/or disorder in the coiled-coil tail. (ii) Both surfaces of undecorated sheets or microtubules, as well as the inner surface of decorated sheets, reveal a strong 4 nm repeat (due to the periodicity of tubulin monomers) and a weak 8 nm repeat (due to slight differences between ␣-and ␤-tubulin). The differences between ␣-and ␤-tubulin on the inner surface are stronger than expected from cryo-electron microscopy of unstained microtubules, indicating the existence of tubulin subdomain-specific surface properties that reflect the surface corrugation and hence metal deposition during evaporation. The 16 nm periodicity visible in some negatively stained specimens (caused by the pairing of cooperatively bound kinesin dimers) is not detected by surface shadowing.

show abstract

Section: Image Analysismentioning

confidence: 99%

Surface Topography of Microtubule Walls Decorated with Monomeric and Dimeric Kinesin Constructs

Hoenger

Doerhoefer²,

Woehlke³

et al. 2000

Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Briefly, square areas were boxed off and their Fourier transforms calculated, from which the lattice parameters were determined. Lattice distortions were determined by locating the cross-correlation between a reference image area with the whole boxed area, and the distortions were corrected by re-interpolation of the image (Henderson et al, 1986). Then, the most probable two-sided plane group was determined by comparing the phase residuals under all possible symmetry constraints (Valpuesta et al, 1994).…”

Section: Electron Microscopy and Image Processingmentioning

confidence: 99%

“…The amplitudes and phases were calculated from the optimized origin, then threefold symmetrized. Finally, an averaged projection map was calculated at 12 Å resolution to include all Fourier terms with measurable signal above noise (IQ Յ 7; Henderson et al, 1986). A negative temperature factor up to 2000 applied in the Fourier synthesis enhanced the contrast toward the higher resolution without introducing spurious features in the map.…”

Section: Electron Microscopy and Image Processingmentioning

confidence: 99%

Structure of TolC, the outer membrane component of the bacterial type I efflux system, derived from two‐dimensional crystals

Koronakis

et al. 1997

Molecular Microbiology

122

View full text Add to dashboard Cite

SummaryTolC is an outer membrane protein required for the export of virulence proteins and toxic compounds without a periplasmic intermediate. We show that TolC is an integral part of the translocator, interacting with inner membrane components, by demonstrating a need for TolC in protein export not only from intact cells but also from sphaeroplasts. To establish the structure of TolC, and thus gain information on how this might be achieved, the protein was purified from the Escherichia coli outer membrane, as a trimer, and crystallized in two-dimensional lattices by reconstitution in phospholipid bilayers. The projection structure at 12 Å resolution showed a threefold symmetric molecule of 58 Å outer diameter, and a single pool of stain filling its centre. Side views parallel to the membrane plane revealed an additional domain outside the membrane. Eighteen membranespanning ␤-strands were predicted for the 51.5 kDa monomer, excluding a 7 kDa C-terminal segment, and this segment was shown to contain a proteinase K-sensitive site that was exposed in reconstituted membranes and sphaeroplasts, but which was protected in intact cells. The combined data suggest that TolC is a trimeric outer membrane protein with each monomer comprising a membrane domain, predicted to be ␤-barrel, and a C-terminal periplasmic domain. The latter could form part of the bridge to the energized inner membrane component of the translocation complex.

show abstract

“…The crystalline structure of a variety of frozen-hydrated biological specimens is preserved to near atomic resolution. Examples include: purple membrane [33,34], catalase [35], crotoxin [36,37], rabbit immunoglobulin FC fragment [38], and T4 DNA helix destabilizing protein [39]. Preparation procedures involving vitrification of the sample also appear to protect noncrystalline biological specimens against the distorting forces of dehydration and ice crystal formation [31].…”

Section: Introductionmentioning

confidence: 99%

Magnification calibration and the determination of spherical virus diameters using cryo-microscopy

Olson

Baker

1989

Ultramicroscopy

View full text Add to dashboard Cite

The diameters of several frozen-hydrated, spherical viruses were determined using polyoma virus as either an external or an internal calibration standard. The methods described provide a reproducible and accurate way to calibrate microscope magnification. The measured diameters are in excellent agreement with respective measurements previously reported for aqueous samples at room temperature using X-ray diffraction methods. These results indicate that the native morphology and dimensions of biological macromolecules are better preserved in frozen-hydrated samples when compared with more conventional electron microscopy techniques such as negative-staining, metal shadowing or thin-sectioning.

show abstract

Structure of purple membrane from halobacterium halobium: recording, measurement and evaluation of electron micrographs at 3.5 Å resolution

Cited by 710 publications

References 35 publications

Surface Topography of Microtubule Walls Decorated with Monomeric and Dimeric Kinesin Constructs

Surface Topography of Microtubule Walls Decorated with Monomeric and Dimeric Kinesin Constructs

Structure of TolC, the outer membrane component of the bacterial type I efflux system, derived from two‐dimensional crystals

Magnification calibration and the determination of spherical virus diameters using cryo-microscopy

Contact Info

Product

Resources

About