Structural stability of vault particles.
Vaults, at 13 MDa, are the largest ribonucleoprotein particles known. In vitro, expression of the major vault protein (MVP) alone in Sf9 insect cells results in the production of recombinant particles with characteristic vault structure. With the ultimate goal of using recombinant vaults as nanocapsules for the delivery of biomolecules, we have employed a variety of spectroscopic techniques (i.e., circular dichroism, fluorescence spectroscopy, and light scattering) along with electron microscopy, to characterize the structural stability of vaults over a wide range of pH (3-8) and temperature (10-90 degrees C). Ten different conformational states of the vaults were identified over the pH and temperature range studied with the most stable region at pH 6-8 below 40 degrees C and least stable at pH 4-6 above 60 degrees C. A unique intermediate molten globulelike state was also identified at pH 6 and approximately 55 degrees C. EM imaging showed the opening of intact vaults into flowerlike structures when transitioning from neutral to acidic pH. This information has potential use in the development of recombinant vaults into nanocapsules for drug delivery since one mechanism by which therapeutic agents entrapped in vaults could be released is through an opening of the intact vault structure.
Esfandiary
Reza
R
Department of Pharmaceutical Chemistry, University of Kansas, 2030 Becker Drive, Lawrence, Kansas 66047, USA.
Kickhoefer
Valerie A
VA
Rome
Leonard H
LH
Joshi
Sangeeta B
SB
Middaugh
C Russell
CR
D000818:Animals; D002460:Cell Line; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004337:Drug Carriers / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D046529:Microscopy, Electron, Transmission; D053769:Nanocapsules / Q000737:chemistry*; D055550:Protein Stability; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000096:biosynthesis / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013696:Temperature; D020394:Vault Ribonucleoprotein Particles / Q000008:administration & dosage / Q000096:biosynthesis / Q000737:chemistry*; D000099187:Major Vault Protein
D004337:Drug Carriers; D053769:Nanocapsules; D011994:Recombinant Proteins; D020394:Vault Ribonucleoprotein Particles; D000099187:Major Vault Protein