Eric R. May

Mechanics of bacteriophage maturation

Capsid maturation with large-scale subunit reorganization occurs in virtually all viruses that use a motor to package nucleic acid into preformed particles. A variety of ensemble studies indicate that the particles gain greater stability during this process, however, it is unknown which material properties of the fragile procapsids change. Using Atomic Force Microscopy-based nano-indentation, we study the development of the mechanical properties during maturation of bacteriophage HK97, a λ-like phage of which the maturation-induced morphological changes are well described. We show that mechanical stabilization and strengthening occurs in three independent ways: (i) an increase of the Young’s modulus, (ii) a strong rise of the capsid’s ultimate strength, and (iii) a growth of the resistance against material fatigue. The Young’s modulus of immature and mature capsids, as determined from thin shell theory, fit with the values calculated using a new multiscale simulation approach. This multiscale calculation shows that the increase in Young’s modulus isn’t dependent on the crosslinking between capsomers. In contrast, the ultimate strength of the capsids does increase even when a limited number of cross-links are formed while full crosslinking appears to protect the shell against material fatigue. Compared to phage λ, the covalent crosslinking at the icosahedral and quasi threefold axes of HK97 yields a mechanically more robust particle than the addition of the gpD protein during maturation of phage λ. These results corroborate the expected increase in capsid stability and strength during maturation, however in an unexpected intricate way, underlining the complex structure of these self-assembling nanocontainers.

Integrin and Defensin Modulate the Mechanical Properties of Adenovirus

ABSTRACT The propensity for capsid disassembly and uncoating of human adenovirus is modulated by interactions with host cell molecules like integrins and alpha defensins. Here, we use atomic force microscopy (AFM) nanoindentation to elucidate, at the single-particle level, the mechanism by which binding of these host molecules affects virus particle elasticity. Our results demonstrate the direct link between integrin or defensin binding and the mechanical properties of the virus. We show that the structure and geometry of adenovirus result in an anisotropic elastic response that relates to icosahedral symmetry. This elastic response changes upon binding host molecules. Whereas integrin binding softens the vertex regions, binding of a human alpha defensin has exactly the opposite effect. Our results reveal that the ability of these host molecules to influence adenovirus disassembly correlates with a direct effect on the elastic strength of the penton region. Host factors that influence adenovirus infectivity thus modulate the elastic properties of the capsid. Our findings reveal a direct link between virus-host interactions and capsid mechanics.

Viral Capsid Equilibrium Dynamics Reveals Nonuniform Elastic Properties

The long wavelength, low-frequency modes of motion are the relevant motions for understanding the continuum mechanical properties of biomolecules. By examining these low-frequency modes, in the context of a spherical harmonic basis set, we identify four elastic moduli that are required to describe the two-dimensional elastic behavior of capsids. This is in contrast to previous modeling and theoretical studies on elastic shells, which use only the two-dimensional Youngs modulus (Y) and the bending modulus (κ) to describe the system. Presumably, the heterogeneity of the structure and the anisotropy of the biomolecular interactions lead to a deviation from the homogeneous, isotropic, linear elastic shell theory. We assign functional relevance of the various moduli governing different deformation modes, including a mode primarily sensed in atomic force microscopy nanoindentation experiments. We have performed our analysis on the T = 3 cowpea chlorotic mottle virus and our estimate for the nanoindentation modulus is in accord with experimental measurements.

Exploring the Symmetry and Mechanism of Virus Capsid Maturation Via an Ensemble of Pathways

Many icosahedral viruses undergo large-scale conformational transitions between icosahedrally symmetric conformations during their life cycles. However, whether icosahedral symmetry is maintained along the transition pathways for this process is unknown. By employing a simplified and directed structure-based potential we compute an ensemble of transition pathways for the maturation transition of bacteriophage HK97. We observe localized symmetry-breaking events, but find that the large-scale displacements are dominated by icosahedrally symmetric deformation modes. We find that all pathways obey a common mechanism characterized by formation of pentameric contacts early in the transition.

On the morphology of viral capsids: elastic properties and buckling transitions.

The morphology of icosahedral viruses ranges from highly spherical to highly faceted, and for some viruses a shape transition occurs during the viral life cycle. This phenomena is predicted from continuum elasticity, via the buckling transition theory by Nelson (Phys. Rev. E 2003, 68, 051910), in which the shape is dependent on the Foppl-von Kármán number (γ), which is a ratio of the two-dimensional Youngs modulus (Y) and the bending modulus (κ). However, until now, no direct calculations have been performed on atomic-level capsid structures to test the predictions of the theory. In this study, we employ a previously described multiscale method by May and Brooks (Phys. Rev. Lett. 2011, 106, 188101) to calculate Y and κ for the bacteriophage HK97, which undergoes a spherical to faceted transition during its viral life cycle. We observe a change in γ consistent with the buckling transition theory and also a significant reduction in κ, which facilitates formation of the faceted state. We go on to examine many capsids from the T = 3 and 7 classes using only elastic network models, which allows us to calculate the ratio Y/κ, without the expense of all-atom molecular dynamics. We observe for the T = 7 capsids, there is strong correlation between the shape of the capsid and γ; however, there is no such correlation for the smaller T = 3 viruses.

pH-induced stability switching of the bacteriophage HK97 maturation pathway.

Many viruses undergo large-scale conformational changes during their life cycles. Blocking the transition from one stage of the life cycle to the next is an attractive strategy for the development of antiviral compounds. In this work, we have constructed an icosahedrally symmetric, low-energy pathway for the maturation transition of bacteriophage HK97. By conducting constant-pH molecular dynamics simulations on this pathway, we identify which residues are contributing most significantly to shifting the stability between the states along the pathway under differing pH conditions. We further analyze these data to establish the connection between critical residues and important structural motifs which undergo reorganization during maturation. We go on to show how DNA packaging can induce spontaneous reorganization of the capsid during maturation.

Molecular modeling of key elastic properties for inhomogeneous lipid bilayers

Fusion and fission of biological membranes play a crucial role in intracellular transport. Until recently, it was believed that membrane shape transformations involved in these processes are driven by proteins. However, recent evidence shows that lipids, by themselves, can drive membrane deformations. It has been hypothesized that the localized formation of certain lipids changes elastic properties of a membrane in such a way that the membrane deforms spontaneously. This study represents a step towards a systematic investigation of the role of various lipids in local changes of membrane elastic properties. We use coarse-grained molecular dynamics (CGMD) simulations to determine possible effects of addition of phosphatidylinositol-4-phosphate (PI4P) lipids on elastic properties of dipalmitoyl phosphatidyl choline (DPPC) lipid bilayers. We investigate the splay (bending) and the molecular tilt moduli of mixed DPPC/PI4P bilayers, as well as the line tension between domains of pure DPPC and mixed DPPC/PI4P bilayers. Although our results indicate negligible effects of PI4P on elastic properties of DPPC bilayers, the developed methodology can be applied to a wide range of lipid systems.

Recent developments in molecular simulation approaches to study spherical virus capsids

Viruses are particularly challenging systems to study via molecular simulation methods. Virus capsids typically consist of over 100 subunit proteins and reach dimensions of over 100 nm; solvated viruses capsid systems can be over 1 million atoms in size. In this review, I will present recent developments which have attempted to overcome the significant computational expense to perform simulations which can inform experimental studies, make useful predictions about biological phenomena and calculate material properties relevant to nanotechnology design efforts.

Evaluation of the hybrid resolution PACE model for the study of folding, insertion, and pore formation of membrane associated peptides

The PACE force field presents an attractive model for conducting molecular dynamics simulations of membrane‐protein systems. PACE is a hybrid model, in which lipids and solvents are coarse‐grained consistent with the MARTINI mapping, while proteins are described by a united atom model. However, given PACE is linked to MARTINI, which is widely used to study membranes, the behavior of proteins interacting with membranes has only been limitedly examined in PACE. In this study, PACE is used to examine the behavior of several peptides in membrane environments, namely WALP peptides, melittin and influenza hemagglutinin fusion peptide (HAfp). Overall, we find PACE provides an improvement over MARTINI for modeling helical peptides, based on the membrane insertion energetics for WALP16 and more realistic melittin pore dynamics. Our studies on HAfp, which forms a helical hairpin structure, do not show the hairpin structure to be stable, which may point toward a deficiency in the model.

Analysis of SecA dimerization in solution.

The Sec pathway mediates translocation of protein across the inner membrane of bacteria. SecA is a motor protein that drives translocation of preprotein through the SecYEG channel. SecA reversibly dimerizes under physiological conditions, but different dimer interfaces have been observed in SecA crystal structures. Here, we have used biophysical approaches to address the nature of the SecA dimer that exists in solution. We have taken advantage of the extreme salt sensitivity of SecA dimerization to compare the rates of hydrogen–deuterium exchange of the monomer and dimer and have analyzed the effects of single-alanine substitutions on dimerization affinity. Our results support the antiparallel dimer arrangement observed in one of the crystal structures of Bacillus subtilis SecA. Additional residues lying within the preprotein binding domain and the C-terminus are also protected from exchange upon dimerization, indicating linkage to a conformational transition of the preprotein binding domain from an open to a closed state. In agreement with this interpretation, normal mode analysis demonstrates that the SecA dimer interface influences the global dynamics of SecA such that dimerization stabilizes the closed conformation.