Award Abstract #0210690
NIRT: The Development of Vault Nano-Capsules

NSF Org:	MCB Division of Molecular and Cellular Biosciences
Initial Amendment Date:	July 31, 2002
Latest Amendment Date:	August 11, 2006
Award Number:	0210690
Award Instrument:	Standard Grant
Program Manager:	John C. Rogers MCB Division of Molecular and Cellular Biosciences BIO Directorate for Biological Sciences
Start Date:	August 1, 2002
Expires:	July 31, 2007 (Estimated)
Awarded Amount to Date:	$1750000
Investigator(s):	Leonard Rome lrome@mednet.ucla.edu (Principal Investigator) Bruce Dunn (Co-Principal Investigator) Harold Monbouquette (Co-Principal Investigator) James Heath (Co-Principal Investigator) Jeffrey Zink (Co-Principal Investigator)
Sponsor:	University of California-Los Angeles 11000 Kinross Avenue LOS ANGELES, CA 90095 310/794-0102
NSF Program(s):	NANOSCALE: INTRDISCPL RESRCH T, PARTICULATE &MULTIPHASE PROCES, BIOCHEMICAL & BIOMASS ENG, PHYSICS-OTHER
Field Application(s):
Program Reference Code(s):	BIOT, AMPP, 9183, 9162, 7202, 1767, 1674, 1132
Program Element Code(s):	1674, 1415, 1402, 1248

ABSTRACT

This Nanoscale Interdisciplinary Research Team (NIRT) project is designed to develop a flexible, targetable nanocapsule by exploiting a naturally occurring nanoscale structure, the vault. Vaults are large (13 MDa) ribonucleoprotein particles composed of multiple copies of three proteins and an RNA, found in nearly all eucaryotic cells. The vault particle is a nanocapsule honed by millions of years of evolution with incredible potential for compound encapsulation, protection, and delivery. The vault nanocapsule can assemble from multiple copies of a few subunits into a stable structure that adheres to and is transported along skeletal networks in the cell, and is likely to open and close in response to cellular signals. Understanding how such a capsule can be manipulated will allow encapsulation of small molecules (drugs, sensors, enzymes, toxins etc.) and targeting the engineered nanostructures to specific tissues, cells, or organelles.

This team plans to modify the vault using a cell-based protein production system and test the concept that vaults can have a broad nanosystems application as malleable nanocapsules. Seven research groups at the University of California, Los Angeles, and other locations (California Institute of Technology; Vanderbilt University), organized by Dr. Leonard Rome, will direct their attention to engineered vault nanocapsules produced in a core production laboratory. Modified vaults will be examined with molecular imaging techniques including negative-stain, cryo-EM single particle reconstruction, atomic force microscopy, and X-ray crystallography. This structural characterization of the vault is an essential first step in designing new vault-based nanocapsules.

Vault nanocapsules will be produced with sequestered metal binding sites and assessed for functional consequences of metal sequestration; for example, cells expressing the engineered particle will be examined to determine whether they have increased resistance to metal toxicity. In other studies, particles will be imbued with functional properties of light emission and magnetic properties to allow the particles to be manipulated in a magnetic field and to probe physiological properties of the vault, i.e., the inflow and outflow of large and small molecules. The ability to manipulate biological nanoparticles in these and other ways offers an opportunity to assemble these particles into structures that may have significant future applications. Spectroscopic techniques will be applied to the engineered particles to allow assessment of the interconversion of opened and closed forms. These studies will lay the groundwork to enable the control of entrapment and release of specific encapsulated materials.

The participants of this team are founding members of the California Nanosystems Institute (CNSI), established to bring together scientists and engineers across disciplines and across institutional boundaries in order to push forward in the area of nanotechnologies. In partnership with the CNSI, the team will establish an educational program designed to advance the field of nanosystems research and technology. This will include training of future researchers and decision-makers and creation of multidisciplinary courses, to enrich school and community resources and influence the technical capabilities of industrial scientists and engineers. In addition, the PI and co-PIs will be involved in research mentoring of undergraduate, graduate and postdoctoral students, curriculum planning, and lecturing. A vault website (www.vaults.arc.ucla.edu) will be maintained and expanded to disseminate information about vaults and vault nanocapsule development. This centralized source of information will include links to published materials and lists of materials and reagents available for sharing.

Funding for this interdisciplinary project is provided through collaborative contributions from the Directorate for Biological Sciences (Division of Molecular and Cellular Biosciences), the Directorate for Engineering (Divisions of Bioengineering & Environmental Systems and Chemical & Transport Systems), and the Directorate for Mathematics & Physical Sciences (Divisions of Physics and Chemistry).

PUBLICATIONS PRODUCED AS A RESULT OF THIS RESEARCH

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76. M.J. Poderycki, V.A. Kickhoefer, C.S. Kaddis, S. Raval-Fernandes, E. Johansson, J.I. Zink, J.A. Loo and L.H. Rome: .  "The Vault Exterior Shell is a Dynamic Structure that Allows Incorporation of Vault-Associated Proteins into its Interior,"  Biochemistry,  v.45,  2006,  p. 12184.

Kickhoefer, V. A. Garcia, Y. Mikyas, Y. Johansson, E. Zhou, J. C. Raval-Fernandes, S. Minoofar, P. Zink, J. I. Dunn, B. Stewart, P. L. Rome, L. H..  "Engineering of vault nanocapsules with enzymatic and fluorescent properties,"  Proc Natl Acad Sci U S A,  v.102,  2005,  p. 4348.

L.E. Goldsmith, M. Yu, L.H. Rome, and H.G. Monbouquette.  "Vault Nanocapsule Dissociation into Halves Triggered at Low pH,"  Biochemistry,  v.46,  2007,  p. 2865.

Mikyas, Y. Makabi, M. Raval-Fernandes, S. Harrington, L. Kickhoefer, V. A. Rome, L. H. Stewart, P. L..  "Cryoelectron microscopy imaging of recombinant and tissue derived vaults: localization of the MVP N termini and VPARP,"  J Mol Biol,  v.344,  2004,  p. 91.


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