Figure 3. Homologs of ORF68 possess similar structures.

(a) Overlay of ORF68 (gray) and BFLF1 (pink) monomers (top) and their homopentameric complexes (bottom). (b) BFLF1 contains at least two zinc fingers, with Zn²⁺ ions shown as a blue-gray sphere. (c) Representative 2D class averages from negative stain EM of UL32. Scale bar = 100 Å. (d) ORF68.stop iSLK cells were lentivirally transcomplemented with plasmids encoding N-terminally Strep-tagged ORF68 or homologs from EBV (BFLF1), MHV68 (mu68), HCMV (UL52), or untagged HSV-1 (UL32). Progeny virion production by these cell lines was assayed by supernatant transfer and flow cytometry of target cells.

Figure 3—figure supplement 1. ConSurf model of ORF68.

Conservation of residues within ORF68 as calculated by the ConSurf server (Ashkenazy et al., 2016), where blue residues are variable (nonconserved) and red are conserved.

Figure 3—figure supplement 2. EM data analysis for BFLF1.

(a) Representative negative stain 2D class averages of BFLF1. Scale bar = 100 Å. (b) Size exclusion chromatography of BFLF1 reveals that it is likely a decamer in solution, rather than a pentamer like ORF68. (c) Representative 2D class averages and data processing workflow for single-particle cryo-EM analyses of BFLF1. The best class was refined to an estimated 3.60 Å resolution, as indicated by the Fourier Shell Correlation (FSC) plot. Scale bar = 100 Å.