Gillian Air  /  Sanjay Bidichandani  /  Robert Broyles
Paul DeAngelis  /  Jay Hanas  /  Martin Levine
Guangpu Li  /  Jialing Lin  /  Hiroyuki Matsumoto  / Blaine Mooers /  Ann Louise Olson 

Karla Rodgers  /  Robert Steinberg  /  Leon Unger  /  Paul Weigel   

Christopher West  /    

Structural basis of the assembly of virus capsids and other multi-protein complexes using X-ray crystallography and solution biophysical techniques.

Our broad goal is to examine the structure and assembly of protein oligomers, determining structures of products, reactants and intermediates, and correlating them with solution studies. A major focus of the lab is to understand the biophysics of virus capsid assembly. The capsid of a spherical of virus is assembled from multiples of 60 protein subunits, arranged with icosahedral symmetry. All projects in the lab involve multimeric viral proteins and enzymes.

Capsid assembly and disassembly are salient events in the virus lifecycle, yet, they are poorly understood and have not been exploited in developing antiviral therapeutics. Virus capsids contain and protect the viral nucleic acid; they may also serve as a delivery system and a metabolic compartment. We are working on two different virus assembly systems Hepatitis B virus (HBV) and Cowpea Chlorotic Mottle virus (CCMV). We are can dissect their assembly reactions by examining assembly and disassembly with a broad range of biophysical and biochemical techniques including fluorescence, light scattering, EM (including image reconstruction), and X-ray crystallography. We have also developed models that describe assembly as a cascade of low order reactions and are generalizable to any virus. Even at this early stage in this research program, we have gained a greater understanding of viral assembly mechanisms.

HBV Assembly: HBV is a DNA virus that causes chronic hepatitis in more than 300 million people, which can lead to cirrhosis or liver cancer. We have examined the assembly of capsids from E. coli expressed dimeric capsid protein. At high pH, we find that HBV capsid assembly begins with slow formation of a trimer of dimers, followed by fast addition of one dimer at a time, till the capsid is complete.  

CCMV Assembly: CCMV is a single stranded RNA virus. It was the first spherical virus assembled in vitro; in vitro assembled virus are infectious and structurally identical to virions purified from plants. CCMV assembly begins with formation of a pentamer of dimers and proceeds by the cooperative addition of dimers. Nucleation and elongation steps have similar rates.

Selected Publications: 

Zlotnick, A., Lee, A., Bourne, C.R., Johnson, J.M., Domanico, P.L., Stray, S.J. (2007) In vitro screening for molecules that affect virus capsid assembly (and other protein association reactions). Nature Protocols 2:490-498. (Cover illustration)

 Zlotnick, A. (2007) Distinguishing reversible from irreversible virus capsid assembly. J. Mol. Biol. 366:14-18.

 Bourne, C. R., Finn, M.G., Zlotnick, A. (2006) Global structural changes in hepatitis B capsids induced by the assembly effector HAP1. J. Virol, 80:11055-11061.

 Stray, S. J., Kopek, B.G., Johnson, J. M., and Zlotnick, A. (2006) A fluorescence-quenching assay identifies molecules that target hepatitis B virus core assembly. Nature Biotech 24:358-362. (Featured in ACS Chemical Biology and in Nature Reviews – Drug Discovery)

 Mukherjee, S., Pfeifer, C., Johnson, J. M., Liu, J., Zlotnick, A.  (2006) Redirecting the Coat Protein of a Spherical Virus to Assemble into Tubular Nanostructures. JACS 128:2538-2539. (Featured in Science News)

 Johnson, J. M., Tang, J., Nyame, Y., Young, M. J., Zlotnick, A.  (2005) Regulating self-assembly of spherical oligomers. Nano Letters 5:765-770.

 Stray, S. J., Bourne, C., Punna, S., Lewis, W. G., Finn, M. G., Zlotnick, A. (2005) A heteroaryldihydropyrimidine enhances and can misdirect assembly of hepatitis B Virus capsid. Proc. Nat. Acad. Sci. USA 102:8138-143.

 Endres, D., Miyahara, M., Moisant, P., Zlotnick, A. (2005) A reaction landscape identifies the intermediates critical for self-assembly of virus capsids and other polyhedral structures. Protein Science 14:1518-1525. (Cover illustration)

 Johnson, J. M., Willits, D. A., Young, M. J., and Zlotnick, A. (2004) Interaction with capsid protein alters RNA structure and the pathway for in vitro assembly of Cowpea Chlorotic Mottle Virus. J. Mol. Biol., 335:455-64.

 Zlotnick, A. (2003). Are Weak Protein-Protein Interactions the General Rule in Capsid Assembly? Virology 315:269-274.

 Ceres, P. and Zlotnick, A. (2002) Hepatitis B Virus Capsid Assembly is Driven by Weak Intersubunit Contacts.  Biochemistry 41:11525-11531.

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