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J Virol. 2016 Oct 14;90(21):9733-9742. doi: 10.1128/JVI.01112-16. Print 2016 Nov 1.

Near-Atomic Resolution Structure of a Highly Neutralizing Fab Bound to Canine Parvovirus.

Author information

1
Department of Medicine, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, USA.
2
Baker Institute for Animal Health, Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, New York, USA.
3
Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA.
4
Department of Medicine, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, USA shafenstein@hmc.psu.edu.

Abstract

Canine parvovirus (CPV) is a highly contagious pathogen that causes severe disease in dogs and wildlife. Previously, a panel of neutralizing monoclonal antibodies (MAb) raised against CPV was characterized. An antibody fragment (Fab) of MAb E was found to neutralize the virus at low molar ratios. Using recent advances in cryo-electron microscopy (cryo-EM), we determined the structure of CPV in complex with Fab E to 4.1 Å resolution, which allowed de novo building of the Fab structure. The footprint identified was significantly different from the footprint obtained previously from models fitted into lower-resolution maps. Using single-chain variable fragments, we tested antibody residues that control capsid binding. The near-atomic structure also revealed that Fab binding had caused capsid destabilization in regions containing key residues conferring receptor binding and tropism, which suggests a mechanism for efficient virus neutralization by antibody. Furthermore, a general technical approach to solving the structures of small molecules is demonstrated, as binding the Fab to the capsid allowed us to determine the 50-kDa Fab structure by cryo-EM.

IMPORTANCE:

Using cryo-electron microscopy and new direct electron detector technology, we have solved the 4 Å resolution structure of a Fab molecule bound to a picornavirus capsid. The Fab induced conformational changes in regions of the virus capsid that control receptor binding. The antibody footprint is markedly different from the previous one identified by using a 12 Å structure. This work emphasizes the need for a high-resolution structure to guide mutational analysis and cautions against relying on older low-resolution structures even though they were interpreted with the best methodology available at the time.

PMID:
27535057
PMCID:
PMC5068525
DOI:
10.1128/JVI.01112-16
[Indexed for MEDLINE]
Free PMC Article

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