Comparison of wild type and genetically engineered nuclear polyhedrosis viruses of Autographa californica for mortality, virus replication and polyhedra production in Trichoplusia ni larvae

Virus replication and polyhedra production of two polyhedron-positive recombinant nuclear polyhedrosis viruses of Autographa californica, AcJHE.KK and AcAaIT which encode juvenile hormone esterase and scorpion toxin, respectively, were compared with those of a plaque purified wild-type nuclear polyhedrosis virus, AcMNPV-C6, in Trichoplusia ni larvae. Though average times required to kill the T. ni larvae increased with the age of the larvae, killing time by either recombinant virus was significantly shorter than that by wild-type virus. Killing time was reduced ca. 30% for AcAaIT-infected larvae and 5 to 8% for AcJHE.KK-infected larvae as compared to that for AcMNPV-C6-infected larvae. The average weight of larvae infected with AcAaIT was significantly lower than that of larvae infected with AcJHE.KK and AcMNPV-C6. The mean numbers of polyhedra produced in each larva inoculated with AcAaIT and AcJHE.KK were ca. 20% and 60%, respectively, of those for AcMNPV-C6. Total virus titers in AcMNPV-C6-infected larvae were significantly higher than those in AcJHE.KK- and AcAaIT-infected larvae until 2 days post infection.     

How baculovirus polyhedra fit square pegs into round holes to robustly package viruses

Natural protein crystals (polyhedra) armour certain viruses, allowing them to survive for years under hostile conditions. We have determined the structure of polyhedra of the baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV), revealing a highly symmetrical covalently cross‐braced robust lattice, the subunits of which possess a flexible adaptor enabling this supra‐molecular assembly to specifically entrap massive baculoviruses. Inter‐subunit chemical switches modulate the controlled release of virus particles in the unusual high pH environment of the target insect's gut. Surprisingly, the polyhedrin subunits are more similar to picornavirus coat proteins than to the polyhedrin of cytoplasmic polyhedrosis virus (CPV). It is, therefore, remarkable that both AcMNPV and CPV polyhedra possess identical crystal lattices and crystal symmetry. This crystalline arrangement must be particularly well suited to the functional requirements of the polyhedra and has been either preserved or re‐selected during evolution. The use of flexible adaptors to generate a powerful system for packaging irregular particles is characteristic of the AcMNPV polyhedrin and may provide a vehicle to sequester a wide range of objects such as biological nano‐particles.     

An Overview of Structural DNA Nanotechnology

Structural DNA Nanotechnology uses unusual DNA motifs to build target shapes and arrangements. These unusual motifs are generated by reciprocal exchange of DNA backbones, leading to branched systems with many strands and multiple helical domains. The motifs may be combined by sticky ended cohesion, involving hydrogen bonding or covalent interactions. Other forms of cohesion involve edge-sharing or paranemic interactions of double helices. A large number of individual species have been developed by this approach, including polyhedral catenanes, a variety of single-stranded knots, and Borromean rings. In addition to these static species, DNA-based nanomechanical devices have been produced that are ultimately targeted to lead to nanorobotics. Many of the key goals of structural DNA nanotechnology entail the use of periodic arrays. A variety of 2D DNA arrays have been produced with tunable features, such as patterns and cavities. DNA molecules have be used successfully in DNA-based computation as molecular representations of Wang tiles, whose self-assembly can be programmed to perform a calculation. About 4 years ago, on the fiftieth anniversary of the double helix, the area appeared to be at the cusp of a truly exciting explosion of applications; this was a correct assessment, and much progress has been made in the intervening period.     

Crystal lattice as biological phenotype for insect viruses

Many insect viruses survive for long periods by occlusion within robust crystalline polyhedra composed primarily of a single polyhedrin protein. We show that two different virus families form polyhedra which, despite lack of sequence similarity in the virally encoded polyhedrin protein, have identical cell constants and a body-centered cubic lattice. It is almost inconceivable that this could have arisen by chance, suggesting that the crystal lattice has been preserved because it is particularly well-suited to its function of packaging and protecting viruses.     

增强蛋白与多角体蛋白共包埋的研究

颗粒体病毒的增强蛋白（enhancin)是一种能显著提高核型多角体病毒（NPV）对昆虫感染力的病毒蛋白。构建了一种不形成多角体但表达粉纹夜蛾颗粒体病毒增强蛋白的重组病毒AcBBH-TnEn,将它与野生型AcMNPV共同感染SF21细胞,经SDS－PAGE、免疫印迹分析、荧光免疫等方法检测证实,增强蛋白与多角体可在同一细胞中同时表达,而且发现所形成的病毒多角体带有增强蛋白。这表明,可以通过混合感染的方式生产带有增强蛋白的病毒多角体。     

Simulating the minimum core for hydrophobic collapse in globular proteins.

To investigate the nature of hydrophobic collapse considered to be the driving force in protein folding, we have simulated aqueous solutions of two model hydrophobic solutes, methane and isobutylene. Using a novel methodology for determining contacts, we can precisely follow hydrophobic aggregation as it proceeds through three stages: dispersed, transition, and collapsed. Theoretical modeling of the cluster formation observed by simulation indicates that this aggregation is cooperative and that the simulations favor the formation of a single cluster midway through the transition stage. This defines a minimum solute hydrophobic core volume. We compare this with protein hydrophobic core volumes determined from solved crystal structures. Our analysis shows that the solute core volume roughly estimates the minimum core size required for independent hydrophobic stabilization of a protein and defines a limiting concentration of nonpolar residues that can cause hydrophobic collapse. These results suggest that the physical forces driving aggregation of hydrophobic molecules in water is indeed responsible for protein folding.     

Evolution of nanoparticle-induced distortion on viral polyhedra

Morphological changes in the polyhedra of the Bombyx mori L. nuclear polyhedrosis virus (BmNPV), a baculovirus causing the deadly grasserie disease in silkworms, brought about by mixing with lipophilically capped amorphous silica nanoparticles (LASN, average size 10 ± 2 nm) were studied with scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy. SEM shows that the regular octagonal polyhedra facets are replaced by a larger number of newly formed irregular ones. The average number of facets reveals a nonlinear growth pattern with nanoparticle (NP) concentration, where an initial linear region ends in a plateau. IR bands corresponding to vibration modes of the capping show (a) a saturation of the area under the band with NP concentration, indicating a correlation with attachment to viral polyhedra and (b) a narrowing of the band per NP from the linear to the plateau portions of the distortion curve, suggesting non-equilibrium and equilibrium situations, respectively.     

Ionic conditions affecting the release and absorption of an alkaline protease associated with the occlusion bodies of insect baculoviruses

Nearly all of the alkaline protease found in the occlusion bodies of baculoviruses (polyhedra for nuclear polyhedrosis and capsules for granulosis viruses) (Baculovirus, subgroup A and B, family Baculoviridae) can be specifically extracted under high ionic concentration. The extraction is directly proportional to the concentrations of NaCl up to 0.25 m. It is not dependent on pH, species of ions, temperature, and incubation time. The protease is reabsorbed under low ionic concentration by protease-extracted and by heat-treated capsules and polyhedra. The protease from Streptomyces griseus is not absorbed. This indicates that the occlusion body proteins have distinct affinity for certain alkaline proteases.     

Fast accumulation of few polyhedra mutants during passage of a Spodoptera frugiperda multicapsid nucleopolyhedrovirus (Baculoviridae) in Sf9 cell cultures

Baculoviruses are a group of viruses that infect invertebrates and that have been used worldwide as a biopesticide against several insect pests of the Order Lepidoptera. In Brazil, the baculovirus Spodoptera frugiperda multicapsid nucleopolyhedrovirus (SfMNPV; Baculoviridae) has been used experimentally to control S. frugiperda (Lepidoptera: Noctuidae), an important insect pest of corn (maize) fields and other crops. Baculoviruses can be produced either in insect larvae or in cell culture bioreactors. A major limitation to the in vitro production of baculoviruses is the rapid generation of mutants when the virus undergoes passages in cell culture. In order to evaluate the potential of in vitro methods of producing SfMNPV on a large-scale, we have multiplied a Brazilian isolate of this virus in cell culture. Extensive formation of few polyhedra mutants was observed after only two passages in Sf9 cells.