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PubMed1990

Two Closely Related Wheat Storage Proteins Follow a Markedly Different Subcellular Route in Xenopus laevis Oocytes.

[alpha]-Gliadins and [gamma]-gliadins are two closely related wheat storage proteins that evolved from a common ancestral gene. However, synthesis of [alpha]-gliadins and [gamma]-gliadins in Xenopus laevis oocytes revealed striking differences in their subcellular routing. The major portion of [alpha]-gliadin accumulated inside the oocyte, whereas most of the [gamma]-gliadin was secreted. Disruption of the Golgi apparatus by monensin revealed that the major part of secretion of [gamma]-gliadin is Golgi mediated. The difference in the subcellular route between [alpha]-gliadin and [gamma]-gliadin may be attributed to differential transport from the endoplasmic reticulum to the Golgi apparatus, a process that is generally the rate-limiting step in protein secretion. Coinjection of the two mRNAs had no effect on their routing, indicating no interaction between them. Our results support the hypothesis that subcellular transport of gliadins in wheat endosperm occurs in two separate routes; one is Golgi mediated, and the other is not. We also show that the subcellular transport may be markedly affected by small structural variations within closely related storage proteins.

Journal Article↗
PubMed2002

Finite element analysis of underwater capacitor micromachined ultrasonic transducers.

A simple electro-mechanical equivalent circuit model is used to predict the behavior of capacitive micromachined ultrasonic transducers (cMUT). Most often, cMUTs are made in silicon and glass plates that are in the 0.5 mm to 1 mm range in thickness. The equivalent circuit model of the cMUT lacks important features such as coupling to the substrate and the ability to predict cross-talk between elements of an array of transducers. To overcome these deficiencies, a flnite element model of the cMUT is constructed using the commercial code ANSYS. Calculation results of the complex load impedance seen by single capacitor cells are presented, then followed by a calculation of the plane wave real load impedance seen by a parallel combination of many cells that are used to make a transducer. Cross-talk between 1-D array elements is found to be due to two main sources: coupling through a Stoneley wave propagating at the transducer-water interface and coupling through Lamb waves propagating in the substrate. To reduce the cross-talk level, the effect of structural variations of the substrate are investigated, which includes a change of its thickness and etched trenches or polymer walls between array elements.

Artifacts↗
PubMed2004

Multislice tomographic imaging and analysis of human breast-equivalent phantoms and biological tissues.

A laser transillumination tomographic system, consisting of electrical, optical, mechanical, and software components, to obtain multislice images of tissue-equivalent breast phantoms and biological tissues, is developed. The tissue-equivalent phantoms are prepared from paraffin wax mixed with wax color pigments by matching their surface backscattered profiles as measured by multiprobe laser reflectometer, with that of respective tissues. The optical parameters of these phantoms are determined by matching their reflectance profiles with that as obtained by Monte Carlo simulation of optical scattering. For multislice tomographic analysis conical breast phantoms of height 80.0 mm and 80.0 mm base diameter with inclusions of different optical properties and dimensions are developed. The resolution of the inclusions in the tomograms depends on their sizes and optical parameters. The minimum size of the inclusion as detected by this procedure in a slice of diameter 50.0 mm is 3.0 mm. The structural variation as observed in the tomograms of phantoms of combination of biological tissues indicates its possible applications in detecting the abnormalities developing in human healthy soft tissues.

Algorithms↗
PubMed2004

Structural and functional analysis of the gpsA gene product of Archaeoglobus fulgidus: a glycerol-3-phosphate dehydrogenase with an unusual NADP+ preference.

NAD(+)-dependent glycerol-3-phosphate dehydrogenase (G3PDH) is generally absent in archaea, because archaea, unlike eukaryotes and eubacteria, utilize glycerol-1-phosphate instead of glycerol-3-phosphate for the biosynthesis of membrane lipids. Surprisingly, the genome of the hyperthermophilic archaeon Archaeoglobus fulgidus comprises a G3PDH ortholog, gpsA, most likely due to horizontal gene transfer from a eubacterial organism. Biochemical characterization proved G3PDH-like activity of the recombinant gpsA gene product. However, unlike other G3PDHs, the up to 85 degrees C thermostable A. fulgidus G3PDH exerted a 15-fold preference for NADPH over NADH. The A. fulgidus G3PDH bears the hallmarks of adaptation to halotolerance and thermophilicity, because its 1.7-A crystal structure showed a high surface density for negative charges and 10 additional intramolecular salt bridges compared to a mesophilic G3PDH structure. Whereas all amino acid residues required for dihydroxyacetone phosphate binding and reductive catalysis are highly conserved, the binding site for the adenine moiety of the NAD(P) cosubstrate shows a structural variation that reflects the observed NADPH preference, for example, by a putative salt bridge between R49 and the 2'-phosphate.

Amino Acid Sequence↗
PubMed1991

Demonstration of initial lymphatics in excised human skin using an extension technique and dye injection.

Using a newly developed technique (extension technique) we succeeded in presenting dermal lymph vessel networks by subepidermal injection of an aqueous Berlin blue solution in excised human skin. Of 94 skin specimens taken from the safety margin of excised melanomas it was possible to demonstrate 32 networks and 14 single dermal lymph vessels. The networks can be documented by macroscopic photographs. Acetic fixation enables us to produce transparent preparations suitable for investigations with a stereo-microscope. The technique described here for macroscopically demonstrating dermal lymph vessels makes it possible to investigate systemically structural variations of lymph vessel networks in human skin.

Acetates↗
PubMed1996

Accordion-like structure of terminal hair follicles of the human scalp.

Terminal hair follicles extracted after ethylenediaminetetraacetic acid treatment preserved the basal cell surface of the outer root sheath well enough to examine the outer surface in detail. Light microscopic observations of whole mount specimens revealed several sawtooth-like structures of follicular epithelium in the lower portion of terminal hair follicle. Vertical sections of the human scalp also showed several invaginations of the outer root sheath of terminal hair follicles in the lower portion, not in association with the arrector pili muscle. Scanning electron microscopy of the same specimens demonstrated accordion-like structures below the sebaceous gland in the lower half of anagen terminal hair follicles and surrounding the entire circumference of the follicles. Sawtooth-like structures were unilaterally observed at the level of the middle or lower portion below the sebaceous gland. With transmission electron microscopy, these structures were seen as undulations of basal cells which preserved hemidesmosomes, although the basal lamina had been peeled off during the extraction procedure. Thus, this accordion-like structure is a genuine structural variation of some human anagen hair follicles and not an artifact.

Basement Membrane↗
PubMed1994

Epitope analysis of isoforms of the major allergen Phl p V by fingerprinting and microsequencing.

The major allergen of timothy grass pollen (Phleum pratense), designated as Phl p V, consists of isoallergenic components of 38 and 32 kDa with pl values of 5.2-7.5 and 4.8-5.9, respectively. The different-sized proteins reveal similarities in IgE reactivity, N-terminal sequence and protein staining. For epitope analysis of these allergens a combination of enzymatic cleavage of electrophoretically separated proteins and immunoblotting techniques with subsequent N-terminal sequencing was performed. After isolation of the components from two-dimensional PAGE gels, proteins were enzymatically cleaved and separated by SDS-PAGE. By endoproteinase Glu-C cleavage six IgE-reactive fragments of each 32 kDa protein and three of each 38 kDa allergen were obtained. Microsequencing of the fragments revealed internal sequences that did not show any similarities between the different-sized allergens. Therefore, we assume only slight structural variations among allergens of similar sizes, whereas the 32 and 38 kDa proteins reveal great differences.

Allergens↗
PubMed2006

History, ocean channels, and distance determine phylogeographic patterns in three widespread Philippine fruit bats (Pteropodidae).

The comparative phylogeography of widespread, codistributed species provides unique insights into regional biodiversity and diversification patterns. I used partial DNA sequences of the mitochondrial genes ND2 and cyt b to investigate phylogeographic structure in three widespread Philippine fruit bats. Ptenochirus jagori is endemic to the oceanic region of the Philippines and is most abundant in lowland primary forest. Macroglossus minimus and Cynopterus brachyotis are most common in disturbed and open habitats and are not endemic. In all three, genetic differentiation is present at multiple spatial scales and is associated to some degree with Pleistocene landbridge island groups. In P. jagori and C. brachyotis, genetic distance is correlated with geographic distance; in C. brachyotis and M. minimus, it is correlated with the sea-crossing distance between islands. P. jagori has the least overall genetic structure of these three species, whereas C. brachyotis and M. minimus have more geographic association among haplotypes, suggesting that phylogeographic patterns are linked to ecology and habitat preference. However, contrary to expectation, the two widespread, disturbed habitat species have more structure than the endemic species. Mismatch distributions suggest rapid changes in effective population size in C. brachyotis and P. jagori, whereas M. minimus appears to be demographically more stable. Geologic and geographic history are important in structuring variation, and phylogeographic patterns are the result of dynamic long-term processes rather than simply reflecting current conditions.

Animals↗
PubMed1992

Interaction of two variable proteins (PilE and PilC) required for pilus-mediated adherence of Neisseria gonorrhoeae to human epithelial cells.

Pili confer the initial ability of Neisseria gonorrhoeae to bind to epithelial cells. Pilin (PilE), the major pilus subunit, and a minor protein termed PilC, reportedly essential for pilus biogenesis, undergo intra-strain phase and structural variation. We demonstrate here that at least two different adherence properties are associated with the gonococcal pili: one is specific for erythrocytes, which is virtually unaffected by PilE variation, and another is specific for epithelial cells, and is modulated in response to the variation of PilE. Based on this finding, mutants of a recA- strain were selected that had lost the ability to bind to human cornea epithelial cells (A-) but retained the ability to form pili (P+) and to agglutinate human erythrocytes (H+). The adherence-negative mutants failed to produce detectable levels of PilC1 or PilC2 proteins, representing piIC phase variants generated in the absence of RecA. The A- pilC phase variants were indistinguishable from their A+ parents and spontaneous A+ revertants with regard to the amount of PilE produced and its electrophoretic mobility, the degrees of piliation and haemagglutination, and the pilE nucleotide sequence. These data demonstrate a central role for PilC in pilus-mediated adherence of N. gonorrhoeae to human epithelial cells and further indicate that neither PilC1 nor PilC2 is obligatory for the assembly of gonococcal pili.

Amino Acid Sequence↗
PubMed1995

The product of the pilQ gene is essential for the biogenesis of type IV pili in Neisseria gonorrhoeae.

The product of the Neisseria gonorrhoeae omc gene possesses regions homologous to those found in members of a protein superfamily that are associated with the translocation of proteins and DNA-protein complexes across the outer membrane. Amongst its protein homologues, Omc has higher overall homology to PilQ, which is required for type IV pilus expression in Pseudomonas aeruginosa, and OrfE, which is required for sequence-specific DNA uptake by Haemophilus influenzae. The function of Omc, however, is unknown and gonococcal omc mutants have not been described. We constructed gonococcal mutants expressing truncated forms of the protein, and found that these mutants are severely defective for both pilus expression and competence for natural transformation. To be consistent with pre-existing pilus gene nomenclature, we have redesignated the gene pilQ instead of omc, and its product, PilQ instead of Omc. The MS11 gene was sequenced and found to differ from the DNA sequence reported for that of another gonococcal strain; these differences were associated with a repeated DNA element, suggesting a genetic basis for structural variation in PilQ. The results also show that PilQ- mutants are distinct from previously described gonococcal pilus-assembly mutants and P. aeruginosa PilQ- mutants by virtue of their expression of rare pilus filaments. Taking these data into account, PilQ is proposed to function in the terminal steps of organelle biogenesis by acting as a pilus channel or pore.

Amino Acid Sequence↗
PubMed2006

The Bacillus subtilis spore coat protein interaction network.

Bacterial spores are surrounded by a morphologically complex, mechanically flexible protein coat, which protects the spore from toxic molecules. The interactions among the over 50 proteins that make up the coat remain poorly understood. We have used cell biological and protein biochemical approaches to identify novel coat proteins in Bacillus subtilis and describe the network of their interactions, in order to understand coat assembly and the molecular basis of its protective functions and mechanical properties. Our analysis characterizes the interactions between 32 coat proteins. This detailed view reveals a complex interaction network. A key feature of the network is the importance of a small subset of proteins that direct the assembly of most of the coat. From an analysis of the network topology, we propose a model in which low-affinity interactions are abundant in the coat and account, to a significant degree, for the coat's mechanical properties as well as structural variation between spores.

Bacillus subtilis↗
PubMed2001

Pressure monitoring of continuous-flow solid-phase peptide synthesis.

We describe the noninvasive real-time pressure monitoring of Boc- and Fmoc-based peptide synthesis. Pressure was measured using a resistance strain gauge attached to the inlet of a continuous-flow reactor of variable volume. In the assembly of the 'difficult' polyalanine sequence, it was shown that pressure monitoring can reveal structural variations of the peptide-resin, e.g. the onset, development and termination of aggregation. This method provided washing minimization that favored substantial saving of solvents. The obtained results demonstrated the advantage of pressure monitoring over swellographic monitoring.

Biochemistry↗
PubMed1985

The structure of the double-stranded RNA pentamer 5'(CACAG) . 5'(CUGUG) determined by nuclear Overhauser enhancement measurements: interproton distance determination and structure refinement on the basis of X-ray coordinates.

The structure in solution of the duplex RNA pentamer 5'(CACAG) . 5'(CUGUG), comprising the stem of the T psi C loop of yeast tRNAPhe, has been investigated by means of one- and two-dimensional nuclear Overhauser enhancement measurements. All non-exchangeable base and sugar proton resonances with the exception of the H5'/H5" sugar resonances are assigned in a sequential manner. From the relative intensities of the cross-peaks obtained in the pure-phase absorption two-dimensional nuclear Overhauser enhancement spectra at several mixing times, it is deduced that the RNA pentamer adopts an A-type conformation in solution. Cross-relaxation rates and interproton distances are determined from the time dependence of the nuclear Overhauser effects, principally by one-dimensional measurements. The structure of the RNA pentamer is then refined by restrained least-squares minimization on the basis of both distance and planarity restraints using fibre diffraction data as an initial model. The refined structure of the RNA pentamer is of the A type but exhibits local structural variations in glycosidic bond and backbone torsion angles as well as in propeller twist, base roll and base tilt angles.

Base Sequence↗
PubMed1995

1H-NMR study of a cobalt-substituted blue copper protein: Pseudomonas aeruginosa Co(II)-azurin.

Substitution of copper by cobalt in blue copper proteins gives a paramagnetic metalloderivative suitable for paramagnetic NMR studies. A thorough analysis of the 1H-NMR spectrum of Pseudomonas aeruginosa Co(II)-azurin is presented here. All the observable contact-shifted signals as well as many other paramagnetic signals from protons placed up to about 1.0 nm around the metal center, including some residues belonging to functionally important parts of the protein like the hydrophobic patch and the His35 region, have been assigned. The results obtained permit the detection and study of structural variations like those originated by the His35 ionization, and allow us to draw a feasible picture of the metal coordination site. Contact-shifted signals correspond to the same five residues which are found in the coordination sphere of the native Cu(II)-azurin, i.e. His46, His117, Cys112, Met121 and Gly45. Among them, the histidine residues present a pattern of resonances typical for histidines coordinated to cobalt in other cobalt protein derivatives, and the cysteine signals clearly indicate a strong interaction with the paramagnetic Co(II) ion. In contrast, the Met121 signals indicate a weak but still existent contact interaction with the metal center. On the other hand, the very weak copper ligand, Gly45, appears here as clearly coordinated to cobalt. Results are consistent with a distorted tetrahedral metal site with the cobalt deviated from the N2S plane towards the Gly45 O axial position and weakly interacting with the Met121 sulfur.

Azurin↗
PubMed1995

The catalytic subunit of cAMP-dependent protein kinase from Ascaris suum. The cloning and structure of a novel subtype of protein kinase A.

A complete cDNA clone encoding the catalytic subunit of cAMP-dependent protein kinase of Ascaris suum was constructed from two overlapping partial clones. The encoded sequence of 337 amino acids is 48% identical with the sequence of mouse C alpha subunit. Approximately the same low similarity was found with the sequence of the C subunit from another nematode, Caenorhabditis elegans. The N-terminal 14 amino acids and the myristoylation site of the mammalian protein are not contained in the enzyme from Ascaris. Two cysteines (Cys33 and Cys319) replace a basic residue in the N-terminal region and an acidic amino acid near the C-terminus which are conserved in all known C subunits from other sources. The substitutions provide the possibility of disulfide bridge formation between the N-terminal and C-terminal parts of the protein. There is strong evidence that a single gene encodes cAMP-dependent protein kinase in Ascaris. Modelling of the sequence into the coordinates of the X-ray structure of the mammalian enzyme suggest a high degree of conservation in the three-dimensional structure. However, structural variations occur at the surface of the protein near the catalytic cleft and are likely to account for the variations in substrate specificity previously observed between the purified protein kinase from Ascaris [Thalhofer, H. P., Daum, G., Harris, B. G. & Hofer, H. W. (1988) J. Biol. Chem. 263, 952-957] and the mammalian enzyme.

Amino Acid Sequence↗
PubMed1998

The comparative anatomical study of the parietal region of the skull of the Korean native goat (Capra hircus).

In the skull of the Korean native goat, the parietal region was classified into four types by the degree of the fusion of the bones, the os interparietale, the os parietale and the squama occipitalis of the os occipitale, and the structural variations of these fusions. The fusion appeared first in the sutura interparietoparietalis and that of the sutura sagittalis of both ossa parietalia was followed. There was no fusion between the os parietale and the squama occipitalis of the os occipitale. These results suggest that the os interparietale developed independently but fused to the os parietale after birth, and the os parietale were developed as paired bones in prenatal life and then fused together according to age.

Animals↗
PubMed1976

An ultrastructural study on precancerous and cancerous lesions of the glandular stomach in rats induced by N-methyl-N'-nitro-N-nitrosoguanidine.

Electron microscopic study was made on precancerous and cancerous lesions in glandular stomach of rats induced by MNNG. Three types of lesions, were found: regenerative glandular hyperplasia, adenomatous hyperplasia and adenocarcinoma. These were compared with the fine structure of similar lesions produced by X-irradiation in the glandular stomach of mice. The precancerous lesion in the present study demonstrated formation of adenomatous hyperplasia by epithelial cells which included mucoid cells characterized by sparce surface microvilli, a few erratic terminal webs, small number of mucous granules, many small mitochondria, and a bizarre nucleus. The development of these mucoid cells seems to be related with the growth of gastric carcinoma. The ultrastructure of adenocarcinomas revealed many structural variations or abnormalities of cellular differentiation. They were classified into 1) surface and pit mucous cell type, 2) pyloric gland cell type, 3) goblet cell type, 4) paneth cell type, 5) intestinal epithelial cell type, 6) endocrine cell type, 7) oncocyte type, 8) filament-rich cell type, and 9) anaplastic cell type involving intracellular microcyst cells. Filament-rich cells were found in poorly differentiated adenocarcinoma or scirrhous carcinoma and may be derived from metaplasia of tumor cell.

Adenocarcinoma↗
PubMed2006

Lack of association between the incidence of testicular germ cell tumors and Y-chromosome haplogroups in the Japanese population.

BACKGROUND: Despite being relatively uncommon, testicular germ cell tumors (TGCT) are the most common malignant disease in young men. Epidemiological studies concerning patients with testicular cancer indicate that the most of them have poor semen quality or testicular dysgenesis. However, many studies have shown that the Y chromosome harbors many candidate genes responsible for spermatogenesis process and development and maintenance of the germ cells. The Y chromosome is thought to have a relationship with the formation and progression of TGCT. MATERIALS AND METHODS: To verify this relationship, we investigated if there is any correlation between the Y chromosome structural variations presented as different haplogroups and the occurrence of TGCT in the Japanese population. Using combined haplogroups based on typing of three Y chromosome polymorphic binary markers, we analyzed 68 TGCT derived from Japanese patients together with randomly selected 104 unrelated healthy Japanese matched male controls who were confirmed as residents of the same geographic area. RESULTS: Our findings showed a lack of association between the incidence of TGCT and the different Y- chromosome haplogroups in Japanese population. CONCLUSION: We concluded that there are no significant variations in males from different Y chromosome lineages regarding their susceptibility or resistance for developing TGCT. The previously hypothesized role of the Y chromosome in the development of TGCT is still uncertain and needs further verification.

Adult↗