Our research is aimed at understanding molecular mechanisms of cellular machines and assemblies using an integrated approach by combining three-dimensional cryo-electron microscopy (cryoEM), 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. We are using cryoEM methods, combined with large scale all-atoms molecular dynamic simulation and biochemical validation to provide reliable structural models of capsid assembly.
Following fusion of the viral and host membranes, the HIV-1 core is released to the cytoplasm of the host cell and transvers to the cell nucleus, during which many cellular factors, including host dependency factor, CypA, and restriction factors, TRIM5α, TRIMCyp and MxB, interact with viral capsid and interfere its uncoating process. To define the structural and functional effects of host factors on the function of capsid and viral infectivity, we are carrying out structural and functional studies of HIV-1 capsid and host factor complexes using cyroEM methods, combined with mutagenesis and functional assays.
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.
While our efforts are driven by biological questions, we have also contributed significantly to the advancement of novel 3D EM methods and technologies, such as those inspired by the bottlenecks we have had to overcome. 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 cells and tissues at molecular resolutions.