NDLI: mRNA display design of fibronectin-based intrabodies that detect and inhibit severe acute respiratory syndrome coronavirus nucleocapsid protein.

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mRNA display design of fibronectin-based intrabodies that detect and inhibit severe acute respiratory syndrome coronavirus nucleocapsid protein.

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mRNA display design of fibronectin-based intrabodies that detect and inhibit severe acute respiratory syndrome coronavirus nucleocapsid protein.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Roberts, Richard W. Sun, Ren Wong, Elaine Liao, Hsiang-i Hwang, Seungmin Deng, Hongyu Baric, Ralph S. Olson, C. Anders
Description	Author Affiliation: Liao HI ( Department of Molecular and Medical Pharmacology, California Nano System Institute, UCLA, Los Angeles, CA 90095, USA.)
Abstract	The nucleocapsid (N) protein of severe acute respiratory syndrome (SARS) coronavirus plays important roles in both viral replication and modulation of host cell processes. New ligands that target the N protein may thus provide tools to track the protein inside cells, detect interaction hot spots on the protein surface, and discover sites that could be used to develop new anti-SARS therapies. Using mRNA display selection and directed evolution, we designed novel antibody-like protein affinity reagents that target SARS N protein with high affinity and selectivity. Our libraries were based on an 88-residue variant of the 10th fibronectin type III domain from human fibronectin (10Fn3). This selection resulted in eight independent 10Fn3 intrabodies, two that require the N-terminal domain for binding and six that recognize the C terminus, one with Kd=1.7 nM. 10Fn3 intrabodies are well expressed in mammalian cells and are relocalized by N in SARS-infected cells. Seven of the selected intrabodies tested do not perturb cellular function when expressed singly in vivo and inhibit virus replication from 11- to 5900-fold when expressed in cells prior to infection. Targeting two sites on SARS-N simultaneously using two distinct 10Fn3s results in synergistic inhibition of virus replication.
ISSN	00219258
e-ISSN	1083351X
Journal	Journal of Biological Chemistry
Issue Number	26
Volume Number	284
Language	English
Publisher	American Society for Biochemistry and Molecular Biology (United States)
Publisher Date	2009-06-26
Publisher Place	United States
Access Restriction	Open
Subject Keyword	Antibodies, Monoclonal Genetics Immunology Gene Expression Profiling Nucleocapsid Proteins Antagonists & Inhibitors RNA, Messenger SARS Virus Physiology Amino Acid Sequence Animals Cells, Cultured Cercopithecus Aethiops Fibronectins Metabolism Fluorescent Antibody Technique Immunoprecipitation Kidney Cytology Molecular Sequence Data Peptide Library Reverse Transcriptase Polymerase Chain Reaction Sequence Homology, Amino Acid Severe Acute Respiratory Syndrome Vero Cells Virus Replication Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Biochemistry Molecular Biology
Content Type	Text
Resource Type	Article
Subject	Cell Biology Biochemistry Molecular Biology

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