Measurement of the accurate mass of a 50 MDa infectious virus

Abstract

Rationale: Bacteriophage P22 is believed to contain a total of 521 copies of 9 different proteins and a 41,724 base pair genome. Despite its enormous size and complexity, phage P22 can be electrosprayed, and it remains intact in ultra-high vacuum where its molar mass distribution has been measured.

Methods: Phage P22 virions were generated by complementation in Salmonella enterica and purified. They were transferred into 100 mM ammonium acetate and then electrosprayed. The masses of individual virions were determined using charge detection mass spectrometry.

Results: The stoichiometry of the protein components of phage P22 is sufficiently well known that the theoretical molar mass can be determined to within a narrow range. The measured average molar mass of phage P22, 52,180 ± 59 kDa, is consistent with the theoretical molar mass and supports the proposed stoichiometry of the components. The intrinsic width of the phage P22 mass distribution can be accounted for by the distribution of DNA packaged by the headful mechanism.

Conclusions: At over 50 MDa, phage P22 is the largest object with a well-defined molar mass to be analyzed by mass spectrometry. The narrow measured mass distribution indicates that the virions survive the transition into the gas phase intact. Copyright © 2016 John Wiley & Sons, Ltd.

Figure 1

A diagram showing the components of infectious phage P22. Adapted from Ref. . The masses of the components are given in Table 1.

Figure 2

Negative-stain, transmission electron micrograph of phage P22. The scale bar is 100 nm.

Figure 3

Mass spectrum of phage P22 measured with CDMS. The spectrum is a histogram constructed with 100 kDa bins. The inset shows an expanded view of the peak at around 52 MDa. In the inset, the blue points are the measured values and the black line is a Gaussian fit to the experimental data. The black points overlaying the main spectrum are a charge vs mass scatter plot showing that the average charge of the phage is about 490 e.