Huilin Li

Brookhaven National Laboratory
Bldg. 463 - P.O. Box 5000
Upton, NY 11973-5000

Joint appointment at the
Department of Biochemistry and Cell Biology,
Stony Brook University

Recent News

Research Interests

Macromolecular assemblies, such as a microtubule shown to the right, perform many critical functions in a cell. Structural information is required for understanding at the molecular and chemical level how these machines function. However, their large sizes make structure determination by NMR and X-ray crystallography difficult. Single particle cryo-electron microscopy (cryo-EM) is becoming an important tool for studying such protein complexes. Uniquely it can reveal the overall protein structures without them having to be crystallized, and it requires only a miniscule amount of sample. Cryo-EM can also be used for studying the conformational changes of molecular assemblies. Our interests are to further advance the cryo-EM method, and to apply the method to understand how the protein machines work. The major instruments we use are a Jeol-2010F TEM/STEM and a Jeol-1200EX TEM.

Molecular mechanism of eukaryotic DNA replication initiation
Eukaryotic chromosomal replication is an intricate process that requires the coordinated and tightly regulated action of numerous molecular machines. Failure to ensure once only replication initiation per cell cycle can result in uncontrolled proliferation and genomic instability, two hallmarks of tumor genesis. The origin recognition complex (ORC), first discovered in yeast by Bruce Stillman, is a six-subunit protein machine conserved in all eukaryotes. Yeast ORC constitutively binds to and marks the replication origin throughout the cell cycle. Licensing of the DNA replication origin starts when the cell division cycle protein Cdc6p binds to ORC. We investigate the structural basis of the molecular mechanism of the DNA replication initiation process.

The M. tuberculosis proteasome and proteasomal ATPase
Tuberculosis kills 1.5-2 million people globally every year. An effective vaccine or chemotherapy has yet to be developed. Recently, the M. tuberculosis (Mtb) proteasome and Mtb proteasomal ATPase (Mpa) were found to be required for Mtb resistance to killing by a source of nitric oxide (NO). NO is required by the host immune system to control Mtb infections. Proteasome and Mpa appear to protect Mtb against NO by degrading proteins after exposure to NO. Thus, Mpa and the Mtb proteasome may be promising targets for the development of anti-Tb chemotherapeutics. We investigate the structure and inhibition mechanism of Mtb proteasome and its associated ATPase with a combined approach of cryo-EM and X-ray crystallography.

Structural characterization of membrane transporters and membrane enzyme complexes We study by electron microscopy the structures of the following membrane protein complexes:   (a) The PapC usher is an outer membrane protein of uropathogenic E. coli. PapC functions as the secretion channel for pilus subunit and as a platform for pilus assembly.   (b) Gamma-Secretase is an intramembranous aspartyl protease that is required for the processing of many membrane proteins, including Notch and amyloid precursor protein. Gamma-Secretase mediated generation of the amyloid beta-protein in brain regions serving memory and cognition can initiate Alzheimer's disease, so the protease has emerged as a key therapeutic target.   (c) Protein N-glycosylation is catalyzed by an enzyme complex oligosaccharyl transferase (OT). The yeast OT complex is composed of nine subunits, all of which are transmembrane proteins.

Developing a TEM/STEM bi-modal cryo-tomography method for subunit mapping of protein assemblies
We develop a hybrid approach, by taking advantage of ultra-structural visualization capability of the cryo-electron microscopy (cryo-EM) and the heavy metal cluster label detection capability of the scanning transmission electron microscopy (STEM) to achieve simultaneously three-dimensional structural visualization and protein mapping. The targeted protein subunit is first labeled by a universal tag such as the Ni-NTA-gold via interaction with a genetically encoded signature such as 6Xhis before assembled into a protein complex.

Recent Publications

• Harada Y., Li H., Wall J.S., Li H., and Lennarz W.J.
  Structural studies and the assembly of the heptameric post-translational translocon complex.
  J. Biol. Chem., 286(4):2956-2965 (Jan 28, 2011). [Epub 2010 Sep 8].  PubMed
• Phan G., Remaut H., Wang T., Allen W.J., Pirker K.F., Lebedev A., Henderson N.S., Geibel S., Volkan E., Yan J., Kunze M.B., Pinkner J.S., Ford B., Kay C.W., Li H., Hultgren S.J., Thanassi D.G., and Waksman G.
  Crystal structure of the FimD usher bound to its cognate FimC-FimH substrate.
  Nature, 474(7349):49-53 (Jun 2 2011).  PubMed
• Burns K.E., Cerda-Maira F.A., Wang T., Li H., Bishai W.R., and Darwin K.H.
  "Depupylation" of prokaryotic ubiquitin-like protein from mycobacterial proteasome substrates.
  Mol Cell., 239(5):821-827 (Sep 10, 2010).  PubMed
• Li D., Li H., Wang T., Pan H., Lin G., and Li H.
  Structural basis for the assembly and gate closure mechanisms of the Mycobacterium tuberculosis 20S proteasome.
  EMBO J., 29(12):2037-2047 (Jun 16, 2010).   PubMed
• Li H., Zhang J., Vierstra R.D., and Li H.
  Quaternary organization of a phytochrome dimer as revealed by cryoelectron microscopy.
  Proc Natl Acad Sci U S A., 107(24):10872-10877 (Jun 15, 2010).   PubMed
• Lin G., Li D., Chidawanyika T., Nathan C., and Li H.
  Fellutamide B is a potent inhibitor of the Mycobacterium tuberculosis proteasome.
  Arch Biochem Biophys., 501(2):214-220 (Sep 15, 2010).   PubMed
• Qi S., Pang Y., Hu Q., Liu Q., Li H., Zhou Y., He T., Liang Q., Liu Y., Yuan X., Luo G., Li H., Wang J., Yan N., and Shi Y.
  Crystal structure of the Caenorhabditis elegans apoptosome reveals an octameric assembly of CED-4.
  Cell, 141(3):446-457 (Apr 30, 2010).  PubMed
• Sun J., and Li H.
  How to operate a cryo-electron microscope.
  Methods Enzymol., 481:231-249 (2010).  PubMed
• Wang T., Darwin K.H., and Li H.
  Binding-induced folding of prokaryotic ubiquitin-like protein on the Mycobacterium proteasomal ATPase targets substrates for degradation.
  Nat. Struct. Mol. Biol., 17(11):1352-1357 (Nov 2010). [Epub 2010 Oct 17].  PubMed
• Evrin C., Clarke P., Zech J., Lurz R., Sun J., Uhle S., Li H., Stillman B., and Speck C.
  A double-hexameric MCM2-7 complex is loaded onto origin DNA during licensing of eukaryotic DNA replication.
  Proc. Natl. Acad. Sci. U.S.A., 106(48):20240-20245 (2009).  PubMed
• Harada, Y., Li, Hua, Li H., and Lennarz, W. J.
  Oligosaccharyltransferase directly binds to ribosome at a location near the translocon-binding site.
  Proceedings of the National Academy of Sciences USA, 106(17):6945-6949 (2009).  PubMed
• Hayashi, M. K., Tang, C., Verpelli, C., Narayanan, R., Stearns, M. H., Xu, R.-M., Li H., Sala, C., and Hayashi, Y.
  The postsynaptic density proteins Homer and Shank form a polymeric network structure.
  Cell, 137(1):159-171 (2009).  PubMed
• Li H. and Thanassi, D. G.
  Use of a combined cryo-EM and X-ray crystallography approach to reveal molecular details of bacterial pilus assembly by the chaperone/usher pathway.
  Current Opinion in Microbiology, 12(2):326-332 (2009).  PubMed
• Li H., Wolfe, M. S., and Selkoe, D. J.
  Toward structural elucidation of the γ-secretase complex.
  Structure, 17(3):326-334 (March, 2009).  PubMed
• Lin G., Li D., de Carvalho L.P., Deng H., Tao H., Vogt G., Wu K., Schneider J., Chidawanyika T., Warren J.D., Li H.*, Nathan C.*.  (* co-corresponding authors)
  Inhibitors selective for mycobacterial versus human proteasomes.
  Nature, 461(7264):621-626 (2009).  PubMed
• Osenkowski P., Li H., Ye W., Li D., Aeschbach L., Fraering P.C., Wolfe M.S., Selkoe D.J., and Li H.
  Cryoelectron microscopy structure of purified gamma-secretase at 12 Å resolution.
  J. Mol. Biol., 385(2):642-652 Epub 2008 Nov 5. (2009).  PubMed
• Wang T., Li H., Lin G., Tang C., Li D., Nathan C., Darwin K.H., and Li H.
  Structural insights on the Mycobacterium tuberculosis proteasomal ATPase Mpa.
  Structure, 17(10):1377-1385 (2009).  PubMed
• Cacquevel M., Aeschbach L., Osenkowski P., Li D., Ye W., Wolfe M.S., Li H., Selkoe D.J. and Fraering P.C.
  Rapid purification of active γ-secretase, an intramembrane protease implicated in Alzheimer’s disease.
  J. Neurochem., 104(1):210-220 (2008).  PubMed
• Chen Z., Speck C., Wendel P., Tang C., Stillman B. and Li H.
  The architecture of the DNA replication origin recognition complex in Saccharomyces cerevisiae.
  Proceedings of the National Academy of Sciences, 105(30):10326-10331 (2008).  PubMed
• Li Hua, Chavan M., Schindelin H., Lennarz W.J. and Li H.
  Structure of the oligosaccharide transferase complex at 12 Å resolution.
  Structure, 16(3):432-440 (2008).  PubMed
• Remaut H., Tang C., Henderson N.S., Pinkner J.S., Wang T., Hultgren S.J., Thanassi D.G., Waksman G. and Li H.
  Fiber formation across the bacterial outer membrane by the chaperone/usher pathway.
  Cell, 133(4):640-652 (2008).  PubMed

Last Modified: December 1, 2011
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One of ten national laboratories overseen and primarily funded by the Office of Science of the U.S. Department of Energy (DOE), Brookhaven National Laboratory conducts research in the physical, biomedical, and environmental sciences, as well as in energy technologies and national security. Brookhaven Lab also builds and operates major scientific facilities available to university, industry and government researchers. Brookhaven is operated and managed for DOE's Office of Science by Brookhaven Science Associates, a limited-liability company founded by Stony Brook University, the largest academic user of Laboratory facilities, and Battelle, a nonprofit, applied science and technology organization.

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