X-ray structural study of noncrystalline regenerated Bombyx mori silk fibroin

X-ray diffraction measurements of regenerated Bombyx mori silk fibroin were carried out to determine its structural characteristic from an analysis of differential radial distribution functions (DRDFs). The temperature dependence of X-ray diffraction patterns from noncrystalline and crystal structures of regenerated silk fibroin was investigated using a high temperature furnace. Time resolved X-ray diffraction profiles were also obtained to construct kinematical models of structural changes caused by the addition of water. DRDFs, calculated from the experimental data, were compared with the DRDFs simulated on the basis of the Monte Carlo method. In order to model the noncrystalline structures, structural units were assumed to be parts of the crystalline structure of silk and those with appropriate structural defects reported previously. From the comparison of experimental and simulated DRDFs, it was determined that noncrystalline regenerated silk consisted of locally ordered atomic sheets similar to the atomic arrangement in the silk I crystal (Type-I sheets), and the final state of the structural change was noncrystalline, consisting of small crystallites, the structure of which is similar to that of silk II (Type-II crystallites). Time resolved DRDFs were also qualitatively interpreted by both the ordering of Type-I sheets and structural changes from Type-I to Type-II. The formation of the small Type-II crystallites obtained in this study was consistent with the nucleation of silk II by birefringence measurements of silk glands and the spinneret of Bombyx mori silkworm reported previously. X-ray diffraction should be a useful technique to understand the structural characteristics of noncrystalline organic materials.     

Light-harvesting ionic dendrimer porphyrins as new photosensitizers for photodynamic therapy

Photodynamic therapy (PDT) is a promising therapeutic modality for treatment of solid tumors. In this study, third-generation aryl ether dendrimer porphyrins (DPs) with either 32 quaternary ammonium groups (32(+)DPZn) or 32 carboxylic groups (32(-)DPZn) were evaluated as a novel, supramolecular class of photosensitizers for PDT. DPs showed a different cell-association profile depending on the positive or negative charge on the periphery, and both DPs eventually localized in membrane-limited organelles. In contrast, protoporphyrin IX (PIX), which is a hydrophobic and relatively low molecular weight photosensitizer used as a control in this study, diffused through the cytoplasm except the nucleus. Confocal fluorescent imaging using organelle-specific dyes indicated that PIX induced severe photodamage to disrupt membranes and intracellular organelles, including the plasma membrane, mitochondrion, and lysosome. On the other hand, cells treated with DPs kept the characteristic fluorescent pattern of such organelles even after photoirradiation. However, notably 32(+)DPZn achieved remarkably higher (1)O(2)-induced cytotoxicity against LLC cells than PIX. Furthermore, both dendrimer porphyrins had far lower dark toxicity as compared with PIX, demonstrating their highly selective photosensitizing effect in combination with a reduced systemic toxicity.     

Modelling the arms race in avian brood parasitism

In brood parasitism, interactions between a parasite and its host lead to a co-evolutionary process called an arms race, in which evolutionary progress on one side provokes a further response on the other side. The host evolves defensive means to reduce the impact of parasitism, while the parasite evolves means to counter the host's defence. To gain insights into the co-evolutionary process of the arms race, a model is developed and analysed, in which the host's defence and the parasite's counterdefence are assumed to be genetically determined. First, the effect of parasite counterdefence on host defence is analysed. I show that parasite counterdefence can critically affect the establishment of host defence, giving rise to three situations in the equilibrium state: The host shows (1) no defence, (2) an intermediate level of defence or (3) perfect defence. Based on these results, the evolution of parasite counterdefence is considered in connection with host defence. It is suggested that the parasite can evolve counterdefence to a certain degree, but once it has established counterdefence beyond this, the host gives up its defence against parasitism provided the defence entails some cost to perform. Dynamic aspects of selection pressure are crucial for these results. Based on these results, I propose a hypothetical evolutionary sequence in the arms race, along which interactions between the host and parasite proceed.     

Why do all host species not show defense against avian brood parasitism: evolutionary lag or equilibrium?

Avian brood parasitism reduces the reproductive success of hosts and is therefore expected to select for host defenses against parasitism, such as an ability to reject parasitic eggs. Field studies have shown that some hosts recognize and reject parasitism, whereas others do not, and the degree of the defense varies from population to population. One long-standing debate concentrates on the differences in the distribution of host defenses observed in hosts parasitized by the brown-headed cowbird and the common cuckoo. The cowbird's hosts show either few or nearly perfect defenses, whereas the cuckoo's hosts have defenses varying from none to complete, with most falling in between the two extremes. To explore the mechanisms underlying this pattern, I constructed a mathematical model in which host defense is assumed to be genetically determined and analyzed how the host defense is established under parasitic pressure. The model shows that differences in the defense-level distribution can be attributed to the difference in the parasite's breeding strategy, generalized or specialized: hosts parasitized by generalists show perfect, none, or intermediate levels of the defense depending on the host abundances, whereas hosts parasitized by specialists always exhibit either none or intermediate levels of the defense if the parasite lacks counter defenses such as egg mimicry. This result provides a testable explanation for the existence of accepter species of the brown-headed cowbird, which might reconcile the previously conflicting hypotheses.     

Olfactory Associative Learning of the Pupal Parasitoid Pimpla luctuosa Smith (Hymenoptera: Ichneumonidae)

Laboratory experiments were conducted to determine the role of learning in olfactory host searching by the ichneumonid pupal parasitoid, Pimpla luctuosa Smith. Females learned to associate novel odors such as vanilla and strawberry with hosts when they oviposited in at least several hosts with the odors. Repeated experiences of hosts with an odor increased the response of the experienced odor, and females that had experienced host odor seven times responded to the experienced odors 90% of the time. Although the response by females to a learned odor gradually decreased with increasing host-deprivation time, 60% of the females that had experienced host odor 7 days earlier still responded to the experienced odor. Females also learned two separate odors associated with hosts at a time and responded to both odors without a preference for one odor over the other. When trained two separate odors with hosts, females learned the second odor more quickly than the first odor. After females experienced several stings in simulated hosts with the previously learned odor, they ceased to respond to the learned odor, suggesting that repeated unrewarding experiences cause females to cease to respond to the learned odors.     

Effect of gelatin on the cyclic AMP response of primocultured hog thyroid cells to acute thyrotropin stimulation.

The cyclic AMP response of cultured hog thyroid cells to acute thyrotropin stimulation was shown to be under a dual regulatory control by thyrotropin: both positive and negative regulation have been described. When added to the culture medium, gelatin (0.25%) promoted the reorganization of the cells into folicle-like structures, as does thyrotropin. Unlike thyrotropin, gelatin did not induce an increase in intracellular cyclic AMP but enhanced the acute cyclic AMP response to thyrotropin in cells cultured in gelatin-containing medium. When both gelatin and thyrotropin were present, the positive effect of low concentrations of hormone (less than 50 microU/ml) was increased whereas the refractory process observed in the presence of higher concentrations of hormone (greater than 50 microU/ml) was unchanged. These effects of gelatin might be mediated by interaction of the denatured collagen molecules with external proteins of the plasma membrane of thyroid cells.     

Single helical structure of curdlan triacetate

The molecular and crystal structure of curdlan triacetate, acetylated (1 → 3)β-D -glucan, was analyzed by means of an x-ray diffraction technique with the help of the linked-atom least-squares method. Unit cell dimensions are a = b = 11.00(1), c(fiber axis) = 22.91 (9) Å, and γ = 120°. The space group is P6₁. The unit cell contains six chemical repeating units related by 6/I-helical symmetry, which is essentially the same as the backbone conformation of one of the modifications (form I) of curdlan. During the refinement calculation, the terminal methyl in every acetyl moiety was elastically restrained to the trans conformation commonly observed in related oligosaccharide structures. The difference Fourier map, the observed and calculated densities, and the thermogravimetric measurement indicated one water molecule per glucose residue. The water oxygen is linked to two carbonyl oxygens in adjacent molecules by hydrogen bonds. The conformation of the primary acetyl moiety is a (skew, -gauche, trans). So far, no skew conformation was observed for the primary acetyl and hydroxyl moieties except in α, β-panose. In both cases, the unusual eclipsed orientation of the primary group is attributed to the hydrogen bond and this conformation is quite different from that of pachyman triacetate. © 1996 John Wiley & Sons, Inc.     

Structural characterization of the MIT domain from human Vps4b

The microtubule interacting and trafficking (MIT) domain is a small protein module of unknown function that is conserved in proteins of diverse function, such as Vps4, sorting nexin 15 (SNX15), and spastin. One non-synonymous single nucleotide polymorphism was reported, which results in a Ile58-to-Met (I58M) substitution in hVps4b. Here, we have determined the solution structure of the MIT domain isolated from the NH(2)-terminus of human Vps4b, an AAA-ATPase involved in multivesicular body formation. The MIT domain adopts an 'up-and-down' three-helix bundle. Comparison with the sequences of other MIT domains clearly shows that the residues involved in inter-helical contacts are well conserved. The Ile58-to-Met substitution resulted a substantial thermal instability. In addition, we found a shallow crevice between helices A and C that may serve as a protein-binding site. We propose that the MIT domain serves as a putative adaptor domain for the ESCRT-III complex involved in endosomal trafficking.     

The effects of skin pressure by clothing on whole gut transit time and amount of feces

The experiment investigated the effects of skin pressure by clothing on whole gut transit time and amount of feces. Comparisons of transit time and amount of feces were taken between 6 young female adults with and without a girdle. The skin pressure applied by a girdle (mean +/- SD) on participant's waist, abdomen and hip region was 16.6 +/- 7.80 mmHg, 17.1 +/- 4.5 mmHg and 12.6 +/- 5.0 mmHg, respectively. The difference of skin pressure with and without a girdle was about 10 mmHg. During each experimental period of 4 days with and without a girdle, the whole gut transit time and amount of feces were measured. The whole gut transit time of each day was determined as the time between the oral intake of each day's markers and their first appearance in the stool. The whole gut transit times of the markers taken on the first day were significantly prolonged (p < 0.05) and those of the markers taken on the second day tended to be prolonged for participant using a girdle (p < 0.1). The whole gut transit time of the markers taken on the third day could not be compared because they were not excreted into the stool. The cumulative increases of feces on the third day and the fourth day were significantly reduced with a girdle (p < 0.05). These results indicate that the skin pressure by clothing has an effect on whole gut transit time and the amount of feces.     

Analysis of local convergence in NMR structure calculation for RNA by a classification system for nucleic acid structure (CSNA)

We are developing a program system, CSNA, to classify a set of structures into groups sharing similar structural characters. In the present study, CSNA was applied to the analysis of NMR structures obtained by the simulated annealing calculation to elucidate local convergences. A 34-mer RNA, U6-34, having a bulge-out region that is derived from the human U6 snRNA is used as a target molecule in the present study. Although the structure calculation was not converged with the conventional method, it was found by the CSNA analysis that the two stem regions in the molecule were converged well. Furthermore, one strand of the bulge-out region (A7-A11) was found to form a continuously stacked structure in two-thirds of calculated structures. In conclusion, CSNA can be a novel tool to elucidate the local convergence of the NMR structure calculations.