TY - JOUR
T1 - Single particle cryoelectron tomography characterization of the structure and structural variability of poliovirus-receptor-membrane complex at 30 Å resolution
AU - Bostina, Mihnea
AU - Bubeck, Doryen
AU - Schwartz, Cindi
AU - Nicastro, Daniela
AU - Filman, David J.
AU - Hogle, James M.
N1 - Funding Information:
This work was supported by NIH Grants AI20566 (to J.M.H.) and an NSF pre-doctoral fellowship (to D.B.). The Boulder Laboratory for 3-Dimensional Electron Microscopy of Cells was supported by NCRR Grant RR000592 (to Andreas Hoenger).
PY - 2007/11
Y1 - 2007/11
N2 - As a long-term goal we want to use cryoelectron tomography to understand how non-enveloped viruses, such as picornaviruses, enter cells and translocate their genomes across membranes. To this end, we developed new image-processing tools using an in vitro system to model viral interactions with membranes. The complex of poliovirus with its membrane-bound receptors was reconstructed by averaging multiple sub-tomograms, thereby producing three-dimensional maps of surprisingly high-resolution (30 Å). Recognizable images of the complex could be produced by averaging as few as 20 copies. Additionally, model-free reconstructions of free poliovirus particles, clearly showing the major surface features, could be calculated from 60 virions. All calculations were designed to avoid artifacts caused by missing information typical for tomographic data ("missing wedge"). To investigate structural and conformational variability we applied a principal component analysis classification to specific regions. We show that the missing wedge causes a bias in classification, and that this bias can be minimized by supplementation with data from the Fourier transform of the averaged structure. After classifying images of the receptor into groups with high similarity, we were able to see differences in receptor density consistent with the known variability in receptor glycosylation.
AB - As a long-term goal we want to use cryoelectron tomography to understand how non-enveloped viruses, such as picornaviruses, enter cells and translocate their genomes across membranes. To this end, we developed new image-processing tools using an in vitro system to model viral interactions with membranes. The complex of poliovirus with its membrane-bound receptors was reconstructed by averaging multiple sub-tomograms, thereby producing three-dimensional maps of surprisingly high-resolution (30 Å). Recognizable images of the complex could be produced by averaging as few as 20 copies. Additionally, model-free reconstructions of free poliovirus particles, clearly showing the major surface features, could be calculated from 60 virions. All calculations were designed to avoid artifacts caused by missing information typical for tomographic data ("missing wedge"). To investigate structural and conformational variability we applied a principal component analysis classification to specific regions. We show that the missing wedge causes a bias in classification, and that this bias can be minimized by supplementation with data from the Fourier transform of the averaged structure. After classifying images of the receptor into groups with high similarity, we were able to see differences in receptor density consistent with the known variability in receptor glycosylation.
KW - Cell entry
KW - Cryoelectron tomography
KW - Poliovirus
KW - Virus-receptor-membrane complex
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U2 - 10.1016/j.jsb.2007.08.009
DO - 10.1016/j.jsb.2007.08.009
M3 - Article
C2 - 17897840
AN - SCOPUS:35148838173
SN - 1047-8477
VL - 160
SP - 200
EP - 210
JO - Journal of Structural Biology
JF - Journal of Structural Biology
IS - 2
ER -