Crystals of cowpea chlorotic mottle virus (CCMV) that diffract X-rays to 3.1 A resolution were grown in a succinate-PEG solution buffered at pH 3.3. The crystals are in space group P2(1)2(1)2(1) with unit cell dimensions of a = 381.26 A, b = 381.26 A, and c = 408.59 A. Four particles occupy the unit cell, placing a single virion in the crystallographic asymmetric unit. Diffraction intensities measured from 196 films collected at the Cornell High Energy Synchrotron Source accounted for 55% of the theoretically possible data to 3.2 A. Unit cell dimensions and rotation function analyses of the X-ray data revealed that the particles were organized in a pseudo-tetragonal relationship with the pseudo-fourfold axis along the crystal c axis. Analysis of electron micrographs of two-dimensional crystals of CCMV revealed a remarkable similarity between these and planes of particles perpendicular to the crystallographic c axis in the three-dimensional crystal.

The three-dimensional structure of Nudaurelia capensis beta virus (N beta V) was reconstructed to 3.2-nm resolution from images of frozen-hydrated virions. The distinctly icosahedral capsid (approximately 40-nm diameter) contains 240 copies of a single 61-kDa protein subunit arranged with T = 4 lattice symmetry. The outer surface of unstained virions compares remarkably well with that previously observed in negatively stained specimens. Inspection of the density map, volume estimates, and model building experiments indicate that each subunit consists of two distinct domains. The large domain (approximately 40 kDa) has a cylindrical shape, approximately 4-nm diameter by approximately 4-nm high, and associates with two large domains of neighboring subunits to form a Y-shaped trimeric aggregate in the outer capsid surface. Four trimers make up each of the 20 planar faces of the capsid. Small domains (approximately 21 kDa) presumably associate at lower radii (approximately 13-16.5 nm) to form a contiguous, non-spherical shell. A T = 4 model, constructed from 80 trimers of the common beta-barrel core motif (approximately 20 kDa) found in many of the smaller T = 3 and pseudo T = 3 viruses, fits the dimensions and features seen in the N beta V reconstruction, suggesting that the contiguous shell of N beta V may be formed by intersubunit contacts between small domains having that motif. The small (approximately 1800 kDa), ssRNA genome is loosely packed inside the capsid with a low average density.

Soil pH regulates the capacity of soils to store and supply nutrients, and thus contributes substantially to controlling productivity in terrestrial ecosystems. However, soil pH is not an independent regulator of soil fertility-rather, it is ultimately controlled by environmental forcing. In particular, small changes in water balance cause a steep transition from alkaline to acid soils across natural climate gradients. Although the processes governing this threshold in soil pH are well understood, the threshold has not been quantified at the global scale, where the influence of climate may be confounded by the effects of topography and mineralogy. Here we evaluate the global relationship between water balance and soil pH by extracting a spatially random sample (n = 20,000) from an extensive compilation of 60,291 soil pH measurements. We show that there is an abrupt transition from alkaline to acid soil pH that occurs at the point where mean annual precipitation begins to exceed mean annual potential evapotranspiration. We evaluate deviations from this global pattern, showing that they may result from seasonality, climate history, erosion and mineralogy. These results demonstrate that climate creates a nonlinear pattern in soil solution chemistry at the global scale; they also reveal conditions under which soils maintain pH out of equilibrium with modern climate.

Background

Studies of simple RNA animal viruses show that cell attachment, particle destabilization and cell entry are complex processes requiring a level of capsid sophistication that is difficult to achieve with a shell containing only a single gene product. Nodaviruses [such as Flock House virus (FHV)] are an exception. We have previously determined the structure of FHV at 3 A resolution, and now combine this information with data from cryo-electron microscopy in an attempt to clarify the process by which nodaviruses infect animal cells.

Results

A difference map was computed in which electron density at 22 A resolution, derived from the 3.0 A resolution X-ray model of the FHV capsid protein, was subtracted from the electron density derived from the cryo-electron microscopy reconstruction of FHV at 22 A resolution. Comparisons of this density with the X-ray model showed that quasi-equivalent regions of identical polypeptide sequences have markedly different interactions with the bulk RNA density. Previously reported biphasic kinetics of particle maturation and the requirement of subunit cleavage for particle infectivity are consistent with these results.

Conclusions

On the basis of this study we propose a model for nodavirus infection that is conceptually similar to that proposed for poliovirus but differs from it in detail. The constraints of a single protein type in the capsid lead to a noteworthy use of quasi-symmetry not only to control the binding of a 'pocket factor' but also to modulate maturation cleavage and to release a pentameric helical bundle (with genomic RNA attached) that may further interact with the cell membrane.

The binding properties of seven mouse monoclonal antibodies (McAbs) raised against cowpea mosaic virus (CPMV) were characterized by conventional and inhibition enzyme-linked immunosorbent assay (ELISA) technique. McAb binding to CPMV on electron microscope (EM) grids was also assayed with gold-labeled anti-mouse antibodies. Two of the seven McAbs (5B2 and 10B7) were found to bind tighter to CPMV than the others in the inhibition ELISA and the EM assay. F(ab) fragments from both of these McAbs were prepared and complexed with CPMV in solution. Electron micrographs of flash frozen (vitrified) samples of native CPMV and CPMV complexed with F(ab) fragments from McAbs 5B2 and 10B7 as well as IgGs from 5B2 were recorded and reconstructions were computed at 23 A resolution for the CPMV/F(ab) complexes and 30 A resolution for the CPMV/IgG complex. Structures of all three complexes clearly displayed the F(ab) fragments distributed with icosahedral symmetry on the surface of CPMV. The IgG bound in a monodentate fashion with only one F(ab) attached to the virus surface. F(ab) fragments from 5B2 and 10B7 bound to nearly identical positions. The refined 2.8 A X-ray structure of CPMV was used to identify the roughly 30 amino acids covered by the F(ab) fragments. The "footprint" spans a subunit interface and appears spatially similar to antigenic site 3B on poliovirus. In a previous, preliminary report of the CPMV/F(ab) 5B2 complex (Wang et al., 1992, Nature 355, 275-278) the wrong enantiomorph of the reconstruction was chosen. This was corrected and, since the F(ab) binds close to a mirror plane, the change in the footprint was minor.

Crystallographic and molecular biological studies of T = 3 nodaviruses (180 identical subunits in the particle) and T = 4 tetraviruses (240 identical subunits in the particle) have revealed similarity in both the architecture of the particles and the strategy for maturation. The comparative studies provide a novel opportunity to examine an apparent evolution of particle size, from smaller (T = 3) to larger (T = 4), with both particles based on similar subunits. The BBV and FHV nodavirus structures are refined at 2.8 A and 3 A respectively, while the N omega V structure is at 6 A resolution. Nevertheless, the detailed comparisons of the noda and tetravirus X-ray electron density maps show that the same type of switching in subunit twofold contacts is used in the T = 3 and T = 4 capsids, although differences must exist between quasi and icosahedral threefold contacts in the T = 4 particle that have not yet been detected. The analyses of primary and tertiary structures of noda and tetraviruses show that N omega V subunits undergo a post assembly cleavage like that observed in nodaviruses and that the cleaved 76 C-terminal residues remain associated with the particle.