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 first 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.
journal_name
Biophys Jjournal_title
Biophysical journalauthors
Clarke RW,Monnier N,Li H,Zhou D,Browne H,Klenerman Ddoi
10.1529/biophysj.107.106351subject
Has Abstractpub_date
2007-08-15 00:00:00pages
1329-37issue
4eissn
0006-3495issn
1542-0086pii
S0006-3495(07)71390-5journal_volume
93pub_type
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