Most bacterial viruses (bacteriophages or just phages) are highly efficient in their ability to infect their often very specific bacterial hosts In general, only one or a few viral particles are necessary to successfully infect one bacterium. As a consequence there has been a long, unfulfilled hope that phages could be used as antibiotics. The need for alternative antibiotics is becoming ever more acute as common bacterial pathogens are developing resistant mutations, making it urgent to develop alternatives such as phage therapy. In addition, recombinant phages have been suggested as a means for gene therapy and as potential antigens for development of vaccines. Bacteriophage T4 has been a model system to study virus structure and function, having advanced biochemical principles of the assembly of biological complexes and protein-protein interactions. We have extensive experience in both structural and functional studies of phage T4. We now wish to expand this knowledge and make it available for potential medical applications. The plan is to analyze the structure and assembly of the capsid (Specific Aim 1), recognition of the host by the phage fibers (Specific Aim 2), the DNA packaging machine (Specific Aim 3), attachment of the packaging machine to the head (Specific Aim 4), and the mechanism of gene delivery (Specific Aim 5). These studies are directed towards engineering phage T4 for specific medical applications to target specific prokaryotic cells (as an antibiotic) and eukaryotic cells (such as cancer cells) for the delivery of toxins. In addition, we are engineering the surface of T4 to carr multiple copies of an epitope for vaccine development. Our primary tools will be molecular biology, protein chemistry, crystallography and electron microscopy. Molecular biology studies will produce pure samples in sufficient quantity for structural and functional studies. Crystallographic studies will be of protein components, providing three-dimensional information at near atomic resolution. Cryo-electron microscopic reconstructions will provide three-dimensional data on the organization of the protein components within the virus. Combining these techniques will generate pseudo-atomic resolution structures of the virus at different stages of its life cycle for functional interpretation and verification.

Public Health Relevance

Bacteriophage T4 is perhaps the most thoroughly studied phage. We plan to extend the structural and functional knowledge of phage T4 to elucidate the mechanisms of assembly, and further, to translate this knowledge into medical applications. We are developing phage T4 into a biomedical platform, to use it as an antibiotic, as a cancer therapeutic, and for the delivery of genes to targeted cells for gene therapy.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI081726-06A1
Application #
8959762
Study Section
Macromolecular Structure and Function C Study Section (MSFC)
Program Officer
Park, Eun-Chung
Project Start
2009-12-15
Project End
2020-04-30
Budget Start
2015-05-01
Budget End
2016-04-30
Support Year
6
Fiscal Year
2015
Total Cost
$652,575
Indirect Cost
$159,445
Name
Purdue University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
072051394
City
West Lafayette
State
IN
Country
United States
Zip Code
47907
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Tao, Pan; Wu, Xiaorong; Rao, Venigalla (2018) Unexpected evolutionary benefit to phages imparted by bacterial CRISPR-Cas9. Sci Adv 4:eaar4134
Rossmann, Michael G (2018) My happy years of collaboration with Fumio Arisaka. Biophys Rev 10:139-140
Chen, Zhenguo; Sun, Lei; Zhang, Zhihong et al. (2017) Cryo-EM structure of the bacteriophage T4 isometric head at 3.3-Å resolution and its relevance to the assembly of icosahedral viruses. Proc Natl Acad Sci U S A 114:E8184-E8193
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Gao, Song; Zhang, Liang; Rao, Venigalla B (2016) Exclusion of small terminase mediated DNA threading models for genome packaging in bacteriophage T4. Nucleic Acids Res 44:4425-39
Arisaka, Fumio; Yap, Moh Lan; Kanamaru, Shuji et al. (2016) Molecular assembly and structure of the bacteriophage T4 tail. Biophys Rev 8:385-396
Fokine, Andrei; Rossmann, Michael G (2016) Common Evolutionary Origin of Procapsid Proteases, Phage Tail Tubes, and Tubes of Bacterial Type VI Secretion Systems. Structure 24:1928-1935
Yap, Moh Lan; Klose, Thomas; Arisaka, Fumio et al. (2016) Role of bacteriophage T4 baseplate in regulating assembly and infection. Proc Natl Acad Sci U S A 113:2654-9

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