The Morais laboratory uses a combination of structural, biophysical, and biochemical methods to elucidate the structure and function of complex macromolecular assemblies and machines. By integrating results from X-ray crystallography, cryo-electron microscopy, and small angle X-ray scattering with traditional biochemical and biophysical methods, we are able to probe how changes in molecular structure give rise to biological function. By understanding the structure-function relationship at the atomic scale, we hope to apply this knowledge towards the development of therapeutics and the rational design of novel nano-motors and machines. A few of the ongoing projects in the Morais lab are discussed below.
Genome packaging motors in dsDNA viruses
Using an exquisitely coordinated molecular process, dsDNA viruses package their genomes into preformed viral capsids to near-crystalline density. Packaging is driven by a transiently assembled molecular motor that converts energy from ATP hydrolysis into the translocation of DNA, working against large entropic, electrostatic, and DNA bending energies that resist DNA compaction. Indeed, viral DNA packaging motors are among the most powerful biological motors known. The virally-encoded ATPases that power genome encapsidation are a sub-group of a large ATPase family whose members are involved in various DNA-remodeling tasks, including cell division, chromosome segregation, DNA recombination, strand separation, and conjugation. Thus, understanding the mechanisms by which viruses package their DNA will also provide insight into general principles of nucleic acid packaging and partitioning in other biological systems. A goal of our laboratory is to visualize the dynamic conformational changes that accompany packaging to dissect motor function and coordination. This work has the potential to inform rational design of nanoscale molecular motors and anti-viral therapeutics that target the genome packaging step.
Cut-away view of the bacteriophage phi29 dsDNA packaging motor. The first inset shows molecular envelopes of the motor components, the second inset shows the X-ray structure of the ATPase that powers genome packaging.
Effect of viral capsid stability and flexibility on viral pathogenesis
Viral capsids must protect the viral genome until it is time to deliver the viral genome to the cell. The stability of the viral capsid required for genome protection function is at odds with the flexibility necessary for the conformational changes required for cell entry and genome delivery. Hyperstable capsids that cannot undergo conformational transitions are inefficient at initiating infection. Conversely, hyperflexible capsids, prone to undergoing conformational changes too easily, are unable to protect the genome. Thus, to remain infectious, capsids of all viruses must maintain a delicate balance between stability and conformational flexibility. We are investigating how this balance is maintained in non-enveloped reoviruses.
Cryo-EM reconstruction of a reovirus infection intermediate.
Archaeal Viruses and Viral Evolution
We are also interested in virus evolution and the viruses which infect the archea. Hyperthermophilic archea possess metabolisms well-suited for the hot anaerobic conditions thought to prevail on early earth, and thus these viruses likely played an important role in the early stages of evolution. Studies on genome organization, replication and regulation of gene expression indicate an evolutionary relationship between archeal viruses and viruses of mesophilic bacteria and eukaryotes. Verification of this hypothesis by sequence comparison is difficult because the rapid evolution of viral genes precludes detection of relationships over large evolutionary distances. However, structural similarity often persists during evolution in spite of vanishing sequence homology. Thus, the structures of viruses infecting hyperthermophilic archaea may provide insights into virus origin and the evolution of viruses and cells. Furthermore, archaeal viruses often have unusual morphologies, and thus understanding their structures and assembly pathways will expand our understanding of macromolecular geometry.
Negative-stained TEM image of a field of “rossettes” of the hyperthermophilic spindle-shaped virus SSV1. A 3D cryo-EM reconstruction of an SSV particle is shown in the lower right.