Supplemental Material for:

Cryoelectron microscopy structure of the adenovirus type 2 temperature sensitive mutant 1 reveals insight into the cell entry defect

Mariena Silvestry1, Steffen Lindert1,2, Jason G. Smith3, Oana Maier4, Christopher M. Wiethoff4, Glen R. Nemerow3, Phoebe L. Stewart1*

1Department of Molecular Physiology and Biophysics, VanderbiltUniversityMedicalCenter, 2215 Garland Avenue, Nashville, TN37232

2Department of Chemistry, VanderbiltUniversity, Nashville, TN37212,

3Department of Immunology and Microbial Science, The Scripps Research Institute, 10550 North Torrey Pines Road, IMM-19, La Jolla, California, 92037

4Department of Microbiology and Immunology, Loyola University Chicago Stritch School of Medicine, Loyola University Chicago, Building 105, 2160 South First Avenue, Maywood, Illinois 60153

*Corresponding Author

Supplemental Figure 1. Masking of the capsid-only and core-plus-capsid regions of a particle image.(A) A projection of the Ad2ts1 reconstruction at 10.5Å resolution viewed along a 3-fold symmetry axis is shown to simulate a particle image. Thesignal-to-noise ratio of this projection image is significantly better than that of the cryoEM particle images. (B) The simulated Ad2ts1 particle image with inner and outer radial masks applied to isolate the capsid-only region. (C) The simulated Ad2ts1 particle image with a circular mask applied to isolate the core-plus-capsid region.

Supplemental Figure 2. The secondary structure prediction algorithms Jufo (5, 6), Psipred (2), and Sam (1, 3) were used to obtain a three-state prediction for Ad5 protein VIII. To increase confidence in the secondary structure prediction, a three-state consensus prediction of the three methods was created by averaging the individual predictions(4). Alpha-helical regions are indicated by red “H” symbols, β-strand regions by blue “S” symbols, and random coil regions by green “C” symbols. The two Ad protease cleavage sites are indicated by vertical bars. The consensus sequence indicates an α-helix of ~22 residues in fragment 1, or two closely spaced α-helices of 6 and 15 residues with a gap of 1 residue.

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