Clarke, RW, Monnier, N, Li, HT et al. (3 more authors) (2007) Two-Color Fluorescence Analysis of Individual Virions Determines the Distribution of the Copy Number of Proteins in Herpes Simplex Virus Particles. Biophysical Journal, 93 (4). pp. 1329-1337. ISSN 0006-3495
Abstract
We present a single virion method to determine absolute distributions of copy number in the protein composition of viruses and apply it to herpes simplex virus type 1. Using two-color coincidence fluorescence spectroscopy, we determine the virion- to-virion variability in copy numbers of fluorescently labeled tegument and envelope proteins relative to a capsid protein by analyzing fluorescence intensity ratios for ensembles of individual dual- labeled virions and fitting the resulting histogram of ratios. Using EYFP- tagged capsid protein VP26 as a reference for fluorescence intensity, we are able to calculate the mean and also, for the. rst time to our knowledge, the variation in numbers of gD, VP16, and VP22 tegument. The measurement of the number of glycoprotein D molecules was in good agreement with independent measurements of average numbers of these glycoproteins in bulk virus preparations, validating the method. The accuracy, straightforward data processing, and high throughput of this technique make it widely applicable to the analysis of the molecular composition of large complexes in general, and it is particularly suited to providing insights into virus structure, assembly, and infectivity.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Keywords: | protein; HSV; single-molecule fluorescence; two-color coincidence |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Engineering & Physical Sciences (Leeds) > School of Chemistry (Leeds) > Inorganic Chemistry (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 12 Jul 2016 10:23 |
Last Modified: | 30 Oct 2016 21:57 |
Published Version: | http://dx.doi.org/10.1529/biophysj.107.106351 |
Status: | Published |
Publisher: | Biophysical Society |
Identification Number: | 10.1529/biophysj.107.106351 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:75849 |