Our research is aimed at understanding molecular mechanisms of cellular machines and assemblies using an integrated approach by combining three-dimensional cryo-electron microscopy and tomography (cryoEM and cryoET), with biochemical, biophysical, computational and molecular biology methods. CryoEM is a powerful tool for structure determination of large protein complexes and macromolecular assemblies, and conformational changes to provide structural snapshots along dynamic processes, as well as 3D architecture of normal and disease cells under their native conditions. Current research efforts in our lab are directed to:
Retroviruses, such as human immunodeficiency virus 1 (HIV-1), contain mature conical capsids that enclose the viral RNA genome, enzymes and accessory proteins. The assembly and stability of the viral capsid are critical to the viral replication life cycle. Structures of the building blocks of the assembly were determined to atomic level, the mechanisms of capsid assembly and uncoating, however, remain unclear. Such information is essential for the development of therapeutic drugs that target viral capsid. Furthermore, many host cell defence proteins, including restriction factors Trim5α, TrimCyp and MxB, target the viral capsid at the early stages of infection and potently inhibit virus replication. These restriction factors appear to function through a remarkable capsid pattern sensing ability that specifically recognizes the assembled capsid, but not the individual capsid protein. Using cutting-edage cryoEM/cryoET technologies, we aim to determine the molecular interactions between the viral capsid and host factors that underpin their capsid pattern-sensing capability and ability to inhibit HIV-1 replication.
Bacteria use chemotaxis signaling pathways to monitor their environment and respond appropriately to change. The essential core signaling unit comprises transmembrane receptors, a histidine kinase CheA, and a coupling protein CheW. Remarkably, bacteria accomplish the extraordinary gain and cooperativity in chemotaxis signaling by arranging a few hundred core signaling units into higher order arrays localized at the cell pole. We developed a novel in vitro reconstitution system to generate signaling arrays, and succeeded in obtaining the first structure of the array using cryoET and sub-tomogram averaging. In the future, we aim to determine the precise molecular mechanisms of chemotaxis cooperative signaling using high-resolution cryoEM and cryoET in combination with site-directed mutagenesis and computational modeling. Our long-term goal is to develop plausible molecular models, at atomic resolution, for the entire signaling pathway by assembling structural “snapshots” of the signaling states.
Driven by biological inquiries, we have contributed significantly to the advancement of novel 3DEM methods and technologies. Technology development has been, and continues to be, an integral part of our research, and we are working on a wide spectrum of technical advances that will be essential to realize the promise of imaging molecules in cells and tissues at near-atomic resolutions in situ in their native environment.