Article
Small compounds targeted to subunit interfaces arrest maturation in a nonenveloped, icosahedral animal virus.
Department of Molecular Biology and Center for Integrative Molecular Biosciences, The Scripps Research Institute, La Jolla, CA 92037, USA.
Journal of Virology (impact factor:
5.4).
08/2004;
78(13):7208-16.
DOI:10.1128/JVI.78.13.7208-7216.2004
Source: PubMed
- Citations (21)
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Cited In (0)
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Article: Methods for reconstructing density maps of "single" particles from cryoelectron micrographs to subnanometer resolution.
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ABSTRACT: Recently the resolution attainable in density maps calculated from cryo-electron micrographs of free-standing virus capsids has advanced to resolutions below 1 nm. This represents a significant milestone in that resolutions of this order potentially allow direct visualization of individual elements of protein secondary structure (i.e., alpha-helices), in addition to the shapes and connectivity of subdomains. We describe here a computational strategy for structural analyses at this level of detail: its principal innovation is a procedure for correcting the contrast transfer function of the electron microscope. Also important is the practice of combining data from pairs of differently defocused micrographs to improve the signal-to-noise ratio of the images, thereby allowing more precise determinations of the particles' orientations and origins and contributing to higher resolution reconstructions. These procedures proved instrumental in our analysis of the capsid of hepatitis B virus at 9-A resolution (Conway et al., 1997, Nature 386, 91-94). Finally, we discuss the prospects for achieving comparable resolutions for isolated macromolecular complexes with lower symmetry or no symmetry and for further extension of the resolution.Journal of Structural Biology 01/2000; 128(1):106-18. · 3.41 Impact Factor -
Article: Large-scale, pH-dependent, quaternary structure changes in an RNA virus capsid are reversible in the absence of subunit autoproteolysis.
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ABSTRACT: The assembly and maturation of the coat protein of a T=4, nonenveloped, single-stranded RNA virus, Nudaurelia capensis omega virus (N omega V), was examined by using a recombinant baculovirus expression system. At pH 7.6, the coat protein assembles into a stable particle called the procapsid, which is 450 A in diameter and porous. Lowering the pH to 5.0 leads to a concerted reorganization of the subunits into a 410-A-diameter particle called the capsid, which has no obvious pores. This conformational change is rapid but reversible until slow, autoproteolytic cleavage occurs in at least 15% of the subunits at the lower pH. In this report, we show that expression of subunits with replacement of Asn-570, which is at the cleavage site, with Thr results in assembly of particles with expected morphology but that are cleavage defective. The conformational change from procapsid to capsid is reversible in N570T mutant virus-like particles, in contrast to wild-type particles, which are locked into the capsid conformation after cleavage of the coat protein. The reexpanded procapsids display slightly different properties than the original procapsid, suggesting hysteretic effects. Because of the stability of the procapsid under near-neutral conditions and the reversible properties of the cleavage-defective mutant, N omega V provides an excellent model for the study of pH-induced conformational changes in macromolecular assemblies. Here, we identify the relationship between cleavage and the conformational change and propose a pH-dependent helix-coil transition that may be responsible for the structural rearrangement in N omega V.Journal of Virology 11/2002; 76(19):9972-80. · 5.40 Impact Factor -
Article: What are the dielectric "constants" of proteins and how to validate electrostatic models?
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ABSTRACT: Implicit models for evaluation of electrostatic energies in proteins include dielectric constants that represent effect of the protein environment. Unfortunately, the results obtained by such models are very sensitive to the value used for the dielectric constant. Furthermore, the factors that determine the optimal value of these constants are far from being obvious. This review considers the meaning of the protein dielectric constants and the ways to determine their optimal values. It is pointed out that typical benchmarks for validation of electrostatic models cannot discriminate between consistent and inconsistent models. In particular, the observed pK(a) values of surface groups can be reproduced correctly by models with entirely incorrect physical features. Thus, we introduce a discriminative benchmark that only includes residues whose pK(a) values are shifted significantly from their values in water. We also use the semimacroscopic version of the protein dipole Langevin dipole (PDLD/S) formulation to generate a series of models that move gradually from microscopic to fully macroscopic models. These include the linear response version of the PDLD/S models, Poisson Boltzmann (PB)-type models, and Tanford Kirkwwod (TK)-type models. Using our different models and the discriminative benchmark, we show that the protein dielectric constant, epsilon(p), is not a universal constant but simply a parameter that depends on the model used. It is also shown in agreement with our previous works that epsilon(p) represents the factors that are not considered explicitly. The use of a discriminative benchmark appears to help not only in identifying nonphysical models but also in analyzing effects that are not reproduced in an accurate way by consistent models. These include the effect of water penetration and the effect of the protein reorganization. Finally, we show that the optimal dielectric constant for self-energies is not the optimal constant for charge-charge interactions.Proteins Structure Function and Bioinformatics 10/2001; 44(4):400-17. · 3.39 Impact Factor
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Keywords
1-anilino-8 naphthalene sulfonate
autocatalytic cleavage
covalent attachment
covalent attachment targets MIANS
critical location
image reconstruction
intermediate configuration
large particle converts
large pores
low-pH MIANS product
MIANS-modified particles
pseudoatomic model
quasi-symmetric surface lattice
rigid body refinement
sealed 395-A icosahedrally
smaller particle
subunits
thiol-reactive fluorophore
two forms
viral subunit
