O'Neil, Alison Linsley.
Engineering bacteriophage P22 as a nanomaterial.
Degree: PhD, College of Letters & Science, 2013, Montana State University
The precise architectures of viruses and virus-like particles are highly advantageous in synthetic materials applications. These nano-size compartments are perfectly suited to act as containers of designed cargo. Not only can these nanocontainers be harnessed as active materials, but they can be exploited for examining the effects of in vivo "cell-like" crowding and confinement on the properties of the encapsulated cargo. The high concentration of many different types of mutually volume excluding macromolecules in the cell causes it to be a crowded and confining environment in which to carry out reactions. Herein, the molecular design of the bacteriophage P22 encapsulation system is described and utilized for the synthesis of active nanomaterials and to explore the effect of encapsulation on the entrapped proteins' properties. In the designed system, any gene can be inserted and results in the fusion of the insert to a truncated form of the P22 scaffold protein. This scaffold protein fusion templates the spontaneous in vivo assembly of P22 capsids and also acts as an encapsulation signal. Once encapsulated, we can examine how crowding and confinement affect inter-molecular communication and activity of the cargo molecules. The P22 system is unique in that the capsid morphology can be altered, without losing the encapsulated cargo, resulting in a doubling of the capsid volume. Thus, the encapsulated fusions can be examined at two different internal concentrations. The packaged capsids contain up to 300 copies of fusion and fill more than 24% of the internal volume with the internal concentration of the fusions in the millimolar range. Not only are these fusions densely and efficiently packaged, but they retain their activity. Described herein is the packaging of fluorescent proteins, enzymes, and active peptides. In all cases, it is shown that the enforced proximity via encapsulation greatly affects the fusions activity compared to the fusion free in solution. To expand the utility of the P22 capsid as a nanomaterial, the inherent asymmetry implored by the portal complex has also been exploited. The P22 encapsulation system has proved to be an effective and versatile vehicle for nanomaterials design.
Advisors/Committee Members: Chairperson, Graduate Committee: Trevor Douglas (advisor), Courtney Reichhardt, Benjamin Johnson, Peter E. Prevelige and Trevor Douglas were co-authors of the article, 'Genetically programmed in vivo packaging and controlled release of protein cargo from bacteriophage P22' in the journal 'Angewandte chemie international edition' which is contained within this thesis. (other), Gautam Basu, Peter E. Prevelige and Trevor Douglas were co-authors of the article, 'Co-confinement of fluorescent proteins: spatially enforced communication of GFP and mCherry encapsulated within the P22 capsid' in the journal 'Biomacromolecules' which is contained within this thesis. (other), Peter E. Prevelige and Trevor Douglas were co-authors of the article, 'Encapsulation within the P22 capsid greatly improves the stability of a phosphotriesterase' submitted to the journal 'Advanced Functional Materials' which is contained within this thesis. (other).
Subjects/Keywords: Microencapsulation.; Nanostructured materials.; Bacteriophages.
to Zotero / EndNote / Reference
APA (6th Edition):
O'Neil, A. L. (2013). Engineering bacteriophage P22 as a nanomaterial. (Doctoral Dissertation). Montana State University. Retrieved from https://scholarworks.montana.edu/xmlui/handle/1/2740
Chicago Manual of Style (16th Edition):
O'Neil, Alison Linsley. “Engineering bacteriophage P22 as a nanomaterial.” 2013. Doctoral Dissertation, Montana State University. Accessed August 15, 2020.
MLA Handbook (7th Edition):
O'Neil, Alison Linsley. “Engineering bacteriophage P22 as a nanomaterial.” 2013. Web. 15 Aug 2020.
O'Neil AL. Engineering bacteriophage P22 as a nanomaterial. [Internet] [Doctoral dissertation]. Montana State University; 2013. [cited 2020 Aug 15].
Available from: https://scholarworks.montana.edu/xmlui/handle/1/2740.
Council of Science Editors:
O'Neil AL. Engineering bacteriophage P22 as a nanomaterial. [Doctoral Dissertation]. Montana State University; 2013. Available from: https://scholarworks.montana.edu/xmlui/handle/1/2740