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Biophysical Studies on the RNA Cores of Satellite Tobacco Mosaic Virus

Published on May 1, 2001in Biophysical Journal3.67
· DOI :10.1016/S0006-3495(01)76206-6
John Day10
Estimated H-index: 10
(UCI: University of California, Irvine),
Yuri G. Kuznetsov6
Estimated H-index: 6
(UCI: University of California, Irvine)
+ 2 AuthorsAlexander McPherson53
Estimated H-index: 53
(UCI: University of California, Irvine)
Cite
Abstract
Satellite tobacco mosaic virus (STMV) was probed using a variety of proteases. Consequences of the degradation were analyzed using gel electrophoresis, quasi-elastic light scattering (QELS), and atomic force microscopy (AFM). Proteolysis rates of 30 minutes for complete degradation of the protein capsid, up to many hours, were investigated. With each protease, degradation of virions 17 nm in diameter was shown by QELS to result in particles of 10 nm diameter, which is that of the RNA core observed in the virion by x-ray diffraction analysis. This was verified by direct visualization with atomic force microscopy. Using QELS, it was further shown that freshly prepared RNA cores remain as individual, stable, 10-nm condensed particles for 12 to 24 h. Clusters of particles then formed, followed by very large aggregates of 500 to 1000 nm diameter. AFM showed that the aggregates were composed of groups of the condensed RNA cores and were not due to unfolding of the nucleic acid. No unfolding of the core particles into extended conformation was seen by AFM until the samples were heated well beyond 90 degrees C. Mass spectrometry of RNA core particles revealed the presence of a major polypeptide whose amino acid sequence corresponded to residues 2 through 25 of the coat protein. Amino acids 13 through 25 were previously observed to be in direct contact with the RNA and are presumably protected from protease digestion. Low resolution difference Fourier analyses indicated the courses of the remainders of the amino terminal strands (amino acids 2-12) in intact virions. Any individual strand appears to have several choices of path, which accounts for the observed disorder at high resolution. These positively charged strands, serving as virtual polyamines, engage the helical segments of RNA. The intimate association of amino acid residues 2 through 25 with RNA likely contributes to the stability of the condensed conformation of the nucleic acid cores.
  • References (23)
  • Citations (21)
Cite
References23
Newest
Published on Jan 1, 1999in Journal of Crystal Growth1.57
Yu. G. Kuznetsov19
Estimated H-index: 19
(UCI: University of California, Irvine),
Alexander J. Malkin22
Estimated H-index: 22
(UCI: University of California, Irvine),
Alexander McPherson53
Estimated H-index: 53
(UCI: University of California, Irvine)
Abstract Atomic force microscopy (AFM) has been used to visualize events arising from the formation of intervening metastable phases at the surfaces of macromolecular crystals growing from solution. Crystals investigated were of the proteins canavalin, thaumatin, lipase, xylanase, and catalase, crystals of transfer RNA, and crystals of satellite tobacco mosaic virus. The appearance of aggregates on crystal surfaces was observed. The aggregates we infer to originate from liquid-protein droplets. ...
Published on Dec 1, 1998in Journal of Molecular Biology5.07
Hiro Tsuruta25
Estimated H-index: 25
(Stanford University),
Vijay S. Reddy29
Estimated H-index: 29
(Scripps Research Institute)
+ 1 AuthorsJohn E. Johnson71
Estimated H-index: 71
(Scripps Research Institute)
Abstract Single crystal diffraction data were collected from virus crystals in the resolution range of 270 to 14 A using a synchrotron X-ray source and a small-angle scattering instrument adapted for single crystal measurements. Reflections were measured from single crystals of the capsid of the double-stranded DNA bacteriophage HK97 and synthetic Flock House virus-like particles (sFHV). The quality of the low-resolution measurements was confirmed by excellent scaling statistics for both data se...
Published on Mar 1, 1998in Journal of Molecular Biology5.07
Steven B. Larson13
Estimated H-index: 13
(UCR: University of California, Riverside),
John Day15
Estimated H-index: 15
(UCR: University of California, Riverside)
+ 1 AuthorsAlexander McPherson53
Estimated H-index: 53
(UCR: University of California, Riverside)
Abstract The molecular structure of satellite tobacco mosaic virus (STMV) has been refined to 1.8 A resolution using X-ray diffraction data collected from crystals grown in microgravity. The final R value was 0.179 and R free was 0.184 for 219,086 independent reflections. The final model of the asymmetric unit contained amino acid residues 13 to 159 of a coat protein monomer, 21 nucleotides, a sulfate ion, and 168 water molecules. The nucleotides were visualized as 30 helical segments of nine ba...
Published on May 1, 1997in Biophysical Journal3.67
Kuznetsov YuG1
Estimated H-index: 1
(UCR: University of California, Riverside),
A Malkin18
Estimated H-index: 18
(UCR: University of California, Riverside)
+ 4 AuthorsAlexander McPherson53
Estimated H-index: 53
(UCR: University of California, Riverside)
Atomic force microscopy (AFM) images at the molecular level have been obtained for a number of different protein and virus crystals. They can be utilized in some special cases to obtain information useful to crystal structure analyses by x-ray diffraction. In particular, questions of space group enantiomer, the packing of molecules within a unit cell, the number of molecules per asymmetric unit, and the dispositions of multiple molecules within the asymmetric unit may be resolved. In addition, b...
Published on Jan 1, 1997
Wah Chiu77
Estimated H-index: 77
,
Roger M. Burnett1
Estimated H-index: 1
,
Robert L. Garcea1
Estimated H-index: 1
Roger M. Burnett and Wah Chiu: Introduction. 1: Sherwood Casjens: Principles of Virion Structure, Function and Assembly. 2: Timothy S. Baker and John E. Johnson: Principles of Virus Structure Determination. 3: Judith M. White, et al.: Attachment and Entry of Influenza Virus Into Host Cells: Pivotal Roles of the Hernagglutin. 4: Michael G. Rossman, et al.: Rhinovirus Attachment and Cell Entry. 5: Thomas J. Smith and Anne G. Mosser: Antibody Mediated Neutralization of Picornaviruses. 6: Marie Chow...
Published on Dec 1, 1995in Physics Today3.09
Carlos Bustamante132
Estimated H-index: 132
(HHMI: Howard Hughes Medical Institute),
David Keller25
Estimated H-index: 25
(UNM: University of New Mexico)
Microscopes have played a fundamental role in the development of biology as an experimental science. It was Robert Hooke who, when using a compound microscope in 1655, noticed that thin slices of cork were made up of identical and small self‐contained units, which he called “cells.” The generalization of this observation and its acceptance, though, had to wait until the late 1830s, when German microscopists Matthias Schleiden and Thcodor Schwann—working independently—introduced the “cell theory”...
Malkin Aj1
Estimated H-index: 1
,
McPherson A1
Estimated H-index: 1
Quasi-elastic light scattering (QELS) was used to investigate quantitatively the mechanisms of nucleation, postnucleation growth, and dissolution in ensembles of both crystalline and amorphous aggregates of satellite tobacco mosaic virus (STMV), ferritin, apoferritin and pumpkin seed globulin. At low supersaturation conditions, as described previously for small molecule crystallization, the metastable region was obtained. Under these conditions aggregation took place, but crystallization did not...
Published on Jan 1, 1994in Progress in Biophysics & Molecular Biology2.70
V.J. Morris1
Estimated H-index: 1
(Norwich Research Park)
Published on May 1, 1993in Journal of Molecular Biology5.07
Steven B. Larson13
Estimated H-index: 13
(UCR: University of California, Riverside),
Stanley Koszelak12
Estimated H-index: 12
(UCR: University of California, Riverside)
+ 3 AuthorsAlexander McPherson53
Estimated H-index: 53
(UCR: University of California, Riverside)
Abstract The crystal structure of satellite tobacco mosaic virus (STMV) has been solved by a combination of multiple isomorphous replacement and molecular replacement methods and refined at 2·9 A resolution to a conventional R -factor of 0·215. SMTV, a T = 1 icosahedral virus, is the smallest whose structure has been determined. The coat protein is an eight-stranded "Swiss roll" β-barrel with an amino-terminal strand that extends away from the β-barrel by more than 60 A. This strand is primarily...
Published on Feb 1, 1993in Journal of Crystal Growth1.57
A Malkin18
Estimated H-index: 18
(UCR: University of California, Riverside),
Josephine Cheung1
Estimated H-index: 1
(UCR: University of California, Riverside),
Alexander McPherson53
Estimated H-index: 53
(UCR: University of California, Riverside)
Quasi-elastic light scattering (QELS) was used to investigate homogeneous and heterogeneous nucleation events in the crystallization of satellite tobacco mosaic virus (STMV). Under conditions of relatively moderate supersaturation (σ ≈ 1.00–1.40), the critical nuclear size supportive of stable crystal growth, Rc, was estimated from time dependent size distribution analyses to be in the range of 30 to 7 virus particles respectively. From the same data, the molar interfacial free energy and the ac...
Cited By21
Newest
Published on May 1, 2015in RNA3.95
Rees F. Garmann11
Estimated H-index: 11
(UCLA: University of California, Los Angeles),
Ajaykumar Gopal13
Estimated H-index: 13
(UCLA: University of California, Los Angeles)
+ 3 AuthorsStephen C. Harvey45
Estimated H-index: 45
(Georgia Institute of Technology)
The lifecycle, and therefore the virulence, of single-stranded (ss)-RNA viruses is regulated not only by their particular protein gene products, but also by the secondary and tertiary structure of their genomes. The secondary structure of the entire genomic RNA of satellite tobacco mosaic virus (STMV) was recently determined by selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE). The SHAPE analysis suggested a single highly extended secondary structure with much less branching t...
Steven B. Larson13
Estimated H-index: 13
(UCI: University of California, Irvine),
John Day10
Estimated H-index: 10
(UCI: University of California, Irvine),
Alexander McPherson53
Estimated H-index: 53
(UCI: University of California, Irvine)
Satellite tobacco mosaic virus (STMV) is among the smallest viruses, having 60 identical subunits arranged with T = 1 icosahedral symmetry. Its crystal structure was solved at 290 K and was refined using, in part, crystals grown in microgravity. Electron-density maps revealed nearly 57% of the genomic ssRNA. Using six flash-cooled crystals, diffraction data were recorded to 1.4 A resolution and independent refinements of the STMV model were carried out versus the previous 1.8 A resolution data r...
Published on Jan 1, 2013
Satellite tobacco mosaic virus (STMV) as the name implies is the satellite virus of the well characterized helper Tobacco mosaic virus (TMV). This satellite virus is 18nm in diameter having a T1 symmetry encapsidating a positive sense genomic RNA of 1058nt in size whose 3' 150nt shares 65% homology with that of its Helper virus. Extensive Structural studies on STMV suggested that its coat protein interacts extensively with the stem-loops of the packaged genomic RNA and hence would be unstable to...
Published on Jul 26, 2012in Journal of Physical Chemistry B2.92
Abhishek Singharoy14
Estimated H-index: 14
(IU: Indiana University Bloomington),
Harshad Joshi9
Estimated H-index: 9
(IU: Indiana University Bloomington)
+ 1 AuthorsP. Ortoleva38
Estimated H-index: 38
(IU: Indiana University Bloomington)
Coarse-grained features of macromolecular assemblies are understood via a set of order parameters (OPs) constructed in terms of their all-atom configuration. OPs are shown to be slowly changing in time and capture the large-scale spatial features of macromolecular assemblies. The relationship of these variables to the classic notion of OPs based on symmetry breaking phase transitions is discussed. OPs based on space warping transformations are analyzed in detail as they naturally provide a conne...
Published on Jan 1, 2012
Norma Hernández-Pedro13
Estimated H-index: 13
,
Edgar Rangel-López8
Estimated H-index: 8
+ 1 AuthorsJulio Sotelo22
Estimated H-index: 22
Properties of biological samples, such as DNA, proteins, components of bacterial surfaces and viruses have been studied extensively and provided the driving force for the outstanding progress of detection methods used in cell biology and physiology. Methods like magnetic twisting cytometry, laser-tracking microrheology, magnetic tweezers, the optical stretcher, and various cell indenters; have been used in the study of cell properties, however, imaging resolution has been low. Atomic Force Micro...
Published on Jan 1, 2012in Frontiers in Microbiology4.26
Hirotaka Ode18
Estimated H-index: 18
(NIH: National Institutes of Health),
Masaaki Nakashima5
Estimated H-index: 5
(Nagoya University)
+ 2 AuthorsHironori Sato28
Estimated H-index: 28
(NIH: National Institutes of Health)
Virus replication in the host proceeds by chains of interactions between viral and host proteins. The interactions are deeply influenced by host immune molecules and anti-viral compounds, as well as by mutations in viral proteins. To understand how these interactions proceed mechanically and how they are influenced by mutations, one needs to know the structures and dynamics of the proteins. Molecular dynamics (MD) simulation is a powerful computational method for delineating very motions of prot...
Published on Oct 1, 2011in Progress in Biophysics & Molecular Biology2.70
Harshad Joshi9
Estimated H-index: 9
(IU: Indiana University Bloomington),
Abhishek Singharoy14
Estimated H-index: 14
(IU: Indiana University Bloomington)
+ 2 AuthorsP. Ortoleva38
Estimated H-index: 38
(IU: Indiana University Bloomington)
A multiscale mathematical and computational approach is developed that captures the hierarchical organization of a microbe. It is found that a natural perspective for understanding a microbe is in terms of a hierarchy of variables at various levels of resolution. This hierarchy starts with the N -atom description and terminates with order parameters characterizing a whole microbe. This conceptual framework is used to guide the analysis of the Liouville equation for the probability density of the...
Published on Jan 28, 2011in Journal of Chemical Physics3.00
Abhishek Singharoy14
Estimated H-index: 14
(IU: Indiana University Bloomington),
S. Cheluvaraja5
Estimated H-index: 5
(IU: Indiana University Bloomington),
P. Ortoleva38
Estimated H-index: 38
(IU: Indiana University Bloomington)
Order parameters (OPs) characterizing the nanoscale features of macromolecules are presented. They are generated in a general fashion so that they do not need to be redesigned with each new application. They evolve on time scales much longer than 10−14 s typical for individual atomic collisions/vibrations. The list of OPs can be automatically increased, and completeness can be determined via a correlation analysis. They serve as the basis of a multiscale analysis that starts with the N-atom Liou...
Published on Dec 1, 2010in Nucleic Acids Research11.15
Yuri G. Kuznetsov1
Estimated H-index: 1
(UAH: University of Alabama in Huntsville),
Jeffrey J. Dowell1
Estimated H-index: 1
(UAH: University of Alabama in Huntsville)
+ 2 AuthorsAlexander McPherson2
Estimated H-index: 2
(UAH: University of Alabama in Huntsville)
Agarose gel electrophoresis, circular dichroism and differential scanning calorimetry showed that single-stranded RNA from satellite tobacco mosaic virus transforms from a conformationally ‘closed state’ at 4°C to a more conformationally ‘open state’ at 65°C. The transition is reversible and shows no hysteresis. Atomic force microscopy (AFM) allowed visualization of the two states and indicated that the conformationally ‘closed state’ probably corresponds to the native encapsidated conformation,...
View next paperAtomic Force Microscopy Analysis of Icosahedral Virus RNA