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Title: In vitro binding of anthrax protective antigen on bacteriophage T4 capsid surface through Hoc-capsid interactions: A strategy for efficient display of large full-length proteins

Abstract

An in vitro binding system is described to display large full-length proteins on bacteriophage T4 capsid surface at high density. The phage T4 icosahedral capsid features 155 copies of a nonessential highly antigenic outer capsid protein, Hoc, at the center of each major capsid protein hexon. Gene fusions were engineered to express the 83-kDa protective antigen (PA) from Bacillus anthracis fused to the N-terminus of Hoc and the 130-kDa PA-Hoc protein was expressed in Escherichia coli and purified. The purified PA-Hoc was assembled in vitro on hoc {sup -} phage particles. Binding was specific, stable, and of high affinity. This defined in vitro system allowed manipulation of the copy number of displayed PA and imposed no significant limitation on the size of the displayed antigen. In contrast to in vivo display systems, the in vitro approach allows all the capsid binding sites to be occupied by the 130-kDa PA-Hoc fusion protein. The PA-T4 particles were immunogenic in mice in the absence of an adjuvant, eliciting strong PA-specific antibodies and anthrax lethal toxin neutralizing antibodies. The in vitro display on phage T4 offers a novel platform for potential construction of customized vaccines against anthrax and other infectious diseases.

Authors:
 [1];  [2];  [2];  [1];  [2];  [2];  [3];  [4]
  1. Department of Biology, 103 McCort Ward Hall, Catholic University of America, 620 Michigan Ave., NE, Washington, DC 20064 (United States)
  2. Division of Retrovirology, Walter Reed Army Institute of Research, 503 Robert Grant Avenue, Silver Spring, MD 20910 (United States)
  3. Bacterial Toxins and Therapeutics Section, National Institute of Allergy and Infectious Diseases, NIH, 30 Convent Dr., Bethesda, MD 20892 (United States)
  4. Department of Biology, 103 McCort Ward Hall, Catholic University of America, 620 Michigan Ave., NE, Washington, DC 20064 (United States). E-mail: rao@cua.edu
Publication Date:
OSTI Identifier:
20779456
Resource Type:
Journal Article
Resource Relation:
Journal Name: Virology; Journal Volume: 345; Journal Issue: 1; Other Information: DOI: 10.1016/j.virol.2005.10.037; PII: S0042-6822(05)00671-9; Copyright (c) 2005 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ANTIBODIES; BACILLUS; BACTERIOPHAGES; ESCHERICHIA COLI; IN VITRO; IN VIVO; INFECTIOUS DISEASES; MICE; PROTEINS; TOXINS; VACCINES

Citation Formats

Shivachandra, Sathish B., Rao, Mangala, Janosi, Laszlo, Sathaliyawala, Taheri, Matyas, Gary R., Alving, Carl R., Leppla, Stephen H., and Rao, Venigalla B. In vitro binding of anthrax protective antigen on bacteriophage T4 capsid surface through Hoc-capsid interactions: A strategy for efficient display of large full-length proteins. United States: N. p., 2006. Web. doi:10.1016/J.VIROL.2005.1.
Shivachandra, Sathish B., Rao, Mangala, Janosi, Laszlo, Sathaliyawala, Taheri, Matyas, Gary R., Alving, Carl R., Leppla, Stephen H., & Rao, Venigalla B. In vitro binding of anthrax protective antigen on bacteriophage T4 capsid surface through Hoc-capsid interactions: A strategy for efficient display of large full-length proteins. United States. doi:10.1016/J.VIROL.2005.1.
Shivachandra, Sathish B., Rao, Mangala, Janosi, Laszlo, Sathaliyawala, Taheri, Matyas, Gary R., Alving, Carl R., Leppla, Stephen H., and Rao, Venigalla B. Sun . "In vitro binding of anthrax protective antigen on bacteriophage T4 capsid surface through Hoc-capsid interactions: A strategy for efficient display of large full-length proteins". United States. doi:10.1016/J.VIROL.2005.1.
@article{osti_20779456,
title = {In vitro binding of anthrax protective antigen on bacteriophage T4 capsid surface through Hoc-capsid interactions: A strategy for efficient display of large full-length proteins},
author = {Shivachandra, Sathish B. and Rao, Mangala and Janosi, Laszlo and Sathaliyawala, Taheri and Matyas, Gary R. and Alving, Carl R. and Leppla, Stephen H. and Rao, Venigalla B.},
abstractNote = {An in vitro binding system is described to display large full-length proteins on bacteriophage T4 capsid surface at high density. The phage T4 icosahedral capsid features 155 copies of a nonessential highly antigenic outer capsid protein, Hoc, at the center of each major capsid protein hexon. Gene fusions were engineered to express the 83-kDa protective antigen (PA) from Bacillus anthracis fused to the N-terminus of Hoc and the 130-kDa PA-Hoc protein was expressed in Escherichia coli and purified. The purified PA-Hoc was assembled in vitro on hoc {sup -} phage particles. Binding was specific, stable, and of high affinity. This defined in vitro system allowed manipulation of the copy number of displayed PA and imposed no significant limitation on the size of the displayed antigen. In contrast to in vivo display systems, the in vitro approach allows all the capsid binding sites to be occupied by the 130-kDa PA-Hoc fusion protein. The PA-T4 particles were immunogenic in mice in the absence of an adjuvant, eliciting strong PA-specific antibodies and anthrax lethal toxin neutralizing antibodies. The in vitro display on phage T4 offers a novel platform for potential construction of customized vaccines against anthrax and other infectious diseases.},
doi = {10.1016/J.VIROL.2005.1},
journal = {Virology},
number = 1,
volume = 345,
place = {United States},
year = {Sun Feb 05 00:00:00 EST 2006},
month = {Sun Feb 05 00:00:00 EST 2006}
}