Stiffness and tanning of sclerites

Tensile studies on cuticle of different chitin fibre architecture indicate that the intimate relationship between physical stiffness and the extent of chemical tanning holds for those cuticles which are mechanically isotropic in their surface planes, but not for those which are mechanically anisotropic.     

Hardening of locust sclerites

The tanning process of presumptive solid cuticle was shown to be correlated with both the tensile force at fracture and the tangent modulus. Both of these physical parameters are closely related to the biochemical events associated with the tanning process.     

Morphology of the elytral base sclerites

The elytral base sclerites (= sclerites located at the articular region between the forewing and thorax in Coleoptera) of selected taxa were examined and homologized. Although the elytral base sclerites are highly modified compared to the wing base sclerites of the other neopterans, they can be homologized by using the conservative wing flapping and folding lines as landmarks. A reduction of the first axillary sclerite was identified as a general trend of the elytral base sclerites, although the sclerite usually has a very important function to mediate flight power from the notum to the wing. This result indicates that the functional constraint against the basal sclerites is relaxed because of the lack of an ability to produce flight power by elytra. In contrast, the elytral folding system formed by the basal sclerites is well retained, which probably occurs because proper wing folding is a key for the shelter function of the elytra. The elytral base sclerites apparently contain more homoplasies than the serially homologous hindwing base sclerites of Coleoptera, which suggests that the structure is less useful for higher-level systematics. However, the faster evolutionary rate of the elytral base sclerites suggests there is potential for studying the lower-level phylogeny of Coleoptera.     

Structural investigations on DNA-protamine complexes.

Protamine·DNA complexes in film and in solution have been investigated by means of infrared linear dichroism, ultraviolet circular dichroism, and laser Raman spectroscopy. At high relative humidity and in solution both infrared linear dichroism and ultraviolet circular dichroism indicate the presence of a modified B form of DNA (designated as B* in our other papers^27,28,46). This modified B form is characterized by a change of the orientation of the \documentclass{article}\pagestyle{empty}\begin{document}$ \widehat{OPO} $\end{document} bisector angle by about 4° with respect to the helical axis when compared to the B form of DNA. At decreasing relative humidities the same modified B form is maintained and no structural transitions B → A (or B → C) normally occurring in free DNA were observed. The absence of the A form in these complexes was also confirmed by laser Raman scattering studies of protamine·DNA complexes. On the basis of these results, a model of the protamine·DNA complex is proposed, which suggests that the presence of apolar amino-acid residues, and probably the folding of the polypeptide chain, is responsible for preventing the B-to-A transition; this occurs either by protecting the high-humidity modified B form against dehydration or by steric interference of this protein probably located in one of the DNA grooves.     

Structural investigations of pneumolysin/lipid complexes

Pneumolysin, a virulence factor from the human pathogen Streptococcus pneumoniae, is a water-soluble protein which forms ring-shaped oligomeric structures upon binding to cholesterol-containing lipid membranes. It induces vesicle aggregation, membrane pore formation and withdrawal of lipid material into non-bilayer proteolipid complexes. Solid-state magic angle spinning and wideline static NMR, together with freeze-fracture electron microscopy, are used to characterize the phase changes in fully hydrated cholesterol-containing lipid membranes induced by the addition ofpneumolysin. A structural model for the proteolipid complexes is proposed where a 30-50-meric pneumolysin ring lines the inside of a lipid torus. Cholesterol is found to be essential to the fusogenic action of pneumolysin.     

Structural investigations of pneumolysin/lipid complexes

Pneumolysin, a virulence factor from the human pathogen Streptococcus pneumoniae, is a water-soluble protein which forms ring-shaped oligomeric structures upon binding to cholesterol-containing lipid membranes. It induces vesicle aggregation, membrane pore formation and withdrawal of lipid material into non-bilayer proteolipid complexes. Solid-state magic angle spinning and wideline static NMR, together with freeze-fracture electron microscopy, are used to characterize the phase changes in fully hydrated cholesterol-containing lipid membranes induced by the addition of pneumolysin. A structural model for the proteolipid complexes is proposed where a 30-50-meric pneumolysin ring lines the inside of a lipid torus. Cholesterol is found to be essential to the fusogenic action of pneumolysin.     

Structural investigations of recombinant urokinase growth factor-like domain

Urokinase-type plasminogen activator (uPA) is a serine protease that converts the plasminogen zymogen into the enzymatically active plasmin. uPA is synthesized and secreted as the single-chain molecule (scuPA) composed of an N-terminal domain (GFD) and kringle (KD) and C-terminal proteolytic (PD) domains. Earlier, the structure of ATF (which consists of GFD and KD) was solved by NMR (A. P. Hansen et al. (1994) Biochemistry, 33, 4847–4864) and by X-ray crystallography alone and in a complex with the soluble form of the urokinase receptor (uPAR, CD87) lacking GPI (C. Barinka et al. (2006) J. Mol. Biol., 363, 482–495). According to these data, GFD contains two β-sheet regions oriented perpendicularly to each other. The area in the GFD responsible for binding to uPAR is localized in the flexible Ω-loop, which consists of seven amino acid residues connecting two strings of antiparallel β-sheet. It was shown by site-directed mutagenesis that shortening of the Ω-loop length by one amino acid residue leads to the inability of GFD to bind to uPAR (V. Magdolen et al. (1996) Eur. J. Biochem., 237, 743–751). Here we show that, in contrast to the above-mentioned studies, we found no sign of the β-sheet regions in GFD in our uPA preparations either free or in a complex with uPAR. The GFD seems to be a rather flexible and unstructured domain, demonstrating in spite of its apparent flexibility highly specific interaction with uPAR both in vitro and in cell culture experiments. Circular dichroism, tryptophan fluorescence during thermal denaturation of the protein, and heteronuclear NMR spectroscopy of ¹⁵N/¹³C-labeled ATF both free and in complex with urokinase receptor were used to judge the secondary structure of GFD of uPA.     

Structural investigations on the non-starchy polysaccharides of apples

Stepwise extraction of cell wall materials from apples with water, aqueous ammonium oxalate and dilute alkali yields polysaccharide preparations from which three components have been isolated on further fractionation in sufficiently homogeneous form for more detailed investigation. Structural studies have furnished evidence for (a) a highly branched α-l-arabinofuranan; (b) pectin fractions of the same general type in which linear 4-linked α-d-galacturonan chains carry α-l-arabinofuranan sub-units; and (c) a fucogalactoxyloglucan.     

Structural investigations of an arabinan from grape juice

An arabinan has been isolated from grape juice and purified by chromatography on polyamide and repeated ethanol precipitations. The structural identity of the arabinan was established by enzymatic degradation of the polysaccharide with a purified -L-arabinofuranosidase and methylation analysis. The results obtained suggest that the arabinan consists of an (1 → 5)-linked backbone of L-arabinofuranosyl residues to which sidechains of L-arabinose are attached in the 3-position.     

A special technique for studying haptoral sclerites of monogeneans

The marginal hooks and anchors have been digested from the haptors of the monogeneans Gyrodactylus salaris and Discocotyle sagittata. These structures then can be measured and drawn more accurately than those surrounded by haptoral tissue. The technique also facilitates the study of isolated hooks with SEM. The results of studies using this digestion technique are presented.     

Two-dimensional versus three-dimensional morphometry of monogenoidean sclerites

A new method of three-dimensional (3-D) analysis of sclerotised structures of monogenoids was performed by processing z-series images using 3D-Doctor. Z-series were obtained from Gomori's trichrome-stained specimens of marine and freshwater monogenoids under laser scanning confocal fluorescence microscopy. Measurements obtained from 3-D images were then compared with those from 2-D images taken from both flattened and unflattened specimens. Data comparison demonstrated that 3-D morphometry allowed avoidance of over-estimation due to deformation and the reduction of errors associated with different spatial orientations. Moreover, study of 3-D images permitted observation of morphological details that are not detectable in 2-D representations.     

Structural investigations on the non-starchy polysaccharides of oat bran

Digestion of oat bran with hog pancreatic α-amylase to hydrolyze starch (～50%) results in solubilization of much β-d-glucan (9%) which is the main non-starchy polysaccharide. This soluble β-d-glucan has been shown by methylation analysis and specific enzymic hydrolysis to contain linear chains with (1→3) and (1→4) linkages in the proportions 1 : 2·6. Compositional and linkage analysis studies on the water-insoluble residue have shown the presence of further β-d-glucan (5%) and arabinoxylan (3%), but only traces of cellulose (<0·5%).     

Structural investigations of lipid, polypeptide and protein multilayers

Langmuir-Blodgett multilayers of lipids, polypeptides and proteins have been examined by X-ray diffraction and infrared spectroscopic methods. The complex polymorphism exhibited by multilayers of glycerides and various phospholipids of different chain length mirror those shown in other three-dimensional structures and suggest that multilayers of lipids can be considered as oriented “crystals”. Both the α and β types of hdyrocarbon chain packing are adopted by different classes of lipids in multilayers.Stable multilayers of the synthetic polypeptide poly-γ-benzyl-l-glutamate consist of α-helical rods stacked in an hexagonal array with a rod axis separation of 14.2 Å. Poly-γ-methyl-l-glutamate behaves similarly but little structural information could be derived from potentially non-helical or sheet-like structures formed by other homopolypeptides. The observation of a single, invariant diffraction line at 9.3 Å for multilayers of a number of water-soluble proteins is consistent with the occurrence of extensive structural reorganization (uncoiling, denaturation) at the air-water interface.