Effect of the viral protease on the dynamics of bacteriophage HK97 maturation intermediates characterized by variance analysis of cryo EM particle ensembles

Abstract

Cryo EM structures of maturation-intermediate Prohead I of bacteriophage HK97 with (PhI(Pro+)) and without (PhI(Pro-)) the viral protease packaged have been reported (Veesler et al., 2014). In spite of PhI(Pro+) containing an additional ∼ 100 × 24 kD of protein, the two structures appeared identical although the two particles have substantially different biochemical properties, e.g., PhI(Pro-) is less stable to disassembly conditions such as urea. Here the same cryo EM images are used to characterize the spatial heterogeneity of the particles at 17Å resolution by variance analysis and show that PhI(Pro-) has roughly twice the standard deviation of PhI(Pro+). Furthermore, the greatest differences in standard deviation are present in the region where the δ-domain, not seen in X-ray crystallographic structures or fully seen in cryo EM, is expected to be located. Thus presence of the protease appears to stabilize the δ-domain which the protease will eventually digest.

Reconstruction of Stochastic 3D Signals With Symmetric Statistics From 2D Projection Images Motivated by Cryo-Electron Microscopy

“…One of the first steps of maturation is the digestion by a virally-encoded protease of the so-called δ domain of the 60 × 7 copies of the capsid peptide that together make up the capsid of the bacteriophage. (The δ domain is roughly the region between radii 93 Å and 193 Å [33], [68] where the outter radius of the particle is 254 Å [69, PDB 3QPR]) The experimental images [68] come from the particle, denoted by PhI Pro+ [33], that contains a protease that is defective so that the particle is trapped in the Prohead I step of maturation. The average outer radius of the capsid is 254 Å [69, PDB 3QPR] and the sphere in which the reconstruction is computed has radius 280 Å.…”

“…In the particular case of Ref. [33], about 10% of the particles changed orientation, including 5% with large orientation changes.…”

“…Ref. [33,Supplemental Figure 1] provides some numerical results on the extent to which estimates of θ i change when the covariance V is simultaneously estimated. In the particular case of Ref.…”

“…Using a model similar to the model of the previous paragraph, the mean and covariance of the random vectors c i can be estimated for each class by a third approach which uses a maximum likelihood estimator computed by an expectationmaximization algorithm in which θ i are the nuisance parameters [33]- [39]. The present paper is an extension of these ideas, which are described in more detail in Section II.…”

“…While this approach fits well within the MATLAB software used in Refs. [33]- [38], more efficient approaches may be possible.…”