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PubMed1988

Immunohistochemical application of monoclonal antibodies to reveal the structure and localization of carbohydrate antigens. N-acetyllactosamine-related carbohydrate antigens in human biliary epithelial cells.

With the use of a panel of monoclonal antibodies against structurally related carbohydrate antigens, finely regulated cellular phenotype of the carbohydrate antigens can be characterized. As a model system, the expression of N-acetyllactosamine-related antigens in human biliary epithelial cells was investigated using 16 murine monoclonal antibodies against different carbohydrate epitopes. Human biliary epithelial cells expressed type 1 backbone-related antigens at all synthetic stages. Type 2 backbone and its monofucosylated structures, i.e., Lex and H, were never detected. More complex type 2 chain antigens, i.e., Ley, A and ALey, were apparently expressed. Types 3 and 4 chain antigens were also shown to be expressed. However, the expression of N-acetyllactosamine-related antigen was polymorphic and differed from one individual to another. Furthermore, in spite of the topographical continuity of the biliary tree, biliary epithelial cells were not always equal in the expression of N-acetyllactosamine-related antigens. As a conclusion, not only the localization but also the structure of carbohydrate antigens can be immunohistochemically studied on the tissue sections with the use of well defined monoclonal antibodies.

ABO Blood-Group System↗
PubMed2001

Characterization of the mouse integrin subunit alpha10 gene and comparison with its human homologue. Genomic structure, chromosomal localization and identification of splice variants.

Alpha10beta1 is a collagen-binding integrin expressed by chondrocytes [Camper et al. (1998) J. Biol. Chem. 273: 20383-20389]. In the present study, the mouse alpha10 gene was isolated from a sCos1 SVJ library and the genomic structure and chromosomal localization was determined. The alpha10 gene consists of 30 translated exons spanning a region of approximately 18 kb genomic DNA. The sequences of all exon/intron borders follow the consensus "gt-ag" rule. A transcription start site, determined by primer extension analysis, was located 38 nucleotides upstream of the initiation ATG site. The 5' flanking region of the transcription start site lacked a TATA-box. The first exon contained, in addition to 38 untranslated nucleotides, the ATG translation start site and the major part of the signal peptide. The alpha10 gene was mapped to mouse chromosome 3 by fluorescence in situ hybridization and is the only integrin subunit localized to this chromosome. When we investigated the expression of alpha10 by PCR we found that both mouse and human articular chondrocytes express extracellular splice variants of the alpha10 subunit. In mouse, exon 26 was extended into the intron by 62 nt, generating a truncated alpha10-chain. In human, exon 25 consisted of 114 nt which were alternately spliced in or out.

Alternative Splicing↗
PubMed2004

Comparative population genetic structures and local adaptation of two mutualists.

Similar patterns of dispersal and gene flow between closely associated organisms may promote local adaptation and coevolutionary processes. We compare the genetic structures of the two species of a plant genus (Roridula gorgonias and R. dentata) and their respective obligately associated hemipteran mutualists (Pameridea roridulae and P. marlothi) using allozymes. In addition, we determine whether genetic structure is related to differences in host choice by Pameridea. Allozyme variation was found to be very structured among plant populations but less so among hemipteran populations. Strong genetic structuring among hemipteran populations was only evident when large distances isolated the plant populations on which they live. Although genetic distances among plant populations were correlated with genetic distances among hemipteran populations, genetic distances of both plants and hemipterans were better correlated with geographic distance. Because Roridula and Pameridea have different scales of gene flow, adaptation at the local population level is unlikely. However, the restricted gene flow of both plants and hemipterans could enable adaptation to occur at a regional level. In choice experiments, the hemipteran (Pameridea) has a strong preference for its carnivorous host plant (Roridula) above unrelated host plants. Pameridea also prefers its host species to its closely related sister species. Specialization at the specific level is likely to reinforce cospeciation processes in this mutualism. However, Pameridea does not exhibit intraspecific preferences toward plants from their natal populations above plants from isolated, non-natal populations.

Adaptation, Biological↗
PubMed2004

Transmission electron microscopy and electron diffraction study of the short-range ordering structure of alpha-LiFeO2.

The basic structure of alpha-LiFeO2, lithium iron oxide, is a cubic NaCl-type structure with a lattice constant of 0.42 nm; some short-range ordering characterized by octahedral clusters exists. The local structure of the short-range ordering was investigated by transmission electron microscopy and electron diffraction. A new short-range ordering structure was found in local areas. The local structure has a cubic lattice with a doubled lattice constant. The occupation factors of cations on Wyckoff sites 4(a) and 4(b) are different from those on 24(d) sites, but the stoichiometric composition in cubic clusters is the same as the macroscopic composition. The number of pairs in which iron cations exist in nearest-neighbor sites and next nearest-neighbor sites is reduced in the structure. This means that a magnetic interaction between the iron cations is reduced by cation ordering even without spin ordering at room temperature.

Journal Article↗
PubMed1999

Order matrix analysis of residual dipolar couplings using singular value decomposition.

The measurement of anisotropic spin interactions, such as residual dipolar couplings, in partially ordered solutions can provide valuable information on biomolecular structure. While the information can be used to refine local structure, it can make a unique contribution in determining the relative orientation of remote parts of molecules, which are locally well structured, but poorly connected based on NOE data. Analysis of dipolar couplings in terms of Saupe order matrices provides a concise description of both orientation and motional properties of locally structured fragments in these cases. This paper demonstrates that by using singular value decomposition as a method for calculating the order matrices, principal frames and order parameters can be determined efficiently, even when a very limited set of experimental data is available. Analysis of 1H-15N dipolar couplings, measured in a two-domain fragment of the barley lectin protein, is used to illustrate the computational method.

Anisotropy↗
PubMed1989

[The localization of structurally specific antigens in squamous cell skin cancer using monoclonal antibodies to epidermal antigens].

Immunomorphological studies by indirect immunofluorescent method using monoclonal antibodies to antigens of basal cells, suprabasal cells and basal membrane of the epidermis were carried out in 8 cases of squamous-cell skin cancer. Specific characteristics of localization of structure-specific antigens (as compared with normal human skin) in squamous-cell skin cancer are discussed.

Aged↗
PubMed2004

Mechanisms of strontium and uranium removal from high-level radioactive waste simulant solutions by the sorbent monosodium titanate.

High-level waste (HLW) is a waste associated with the dissolution of spent nuclear fuel for the recovery of weapons-grade material. It is the priority problem for the U.S. Department of Energy's Environmental Management Program. Current HLW treatment processes at the Savannah River Site (Aiken, SC) include the use of monosodium titanate (MST, with a similar stoichiometry to NaTi2O5 x xH2O) to concentrate strontium (Sr) and actinides. The high affinity of MST for Sr and actinides in HLW solutions rich in Na+ is poorly understood. Mechanistic information about the nature of radionuclide uptake will provide insight about MST treatment reliability. Our study characterized the morphology of MST and the chemistry of sorbed Sr2+ and uranium [U(VI)] as uranyl ion, UO2(2+), on MST, which were added (individually) from stock solutions of Sr and 238U(VI) with spectroscopic and transmission electron microscopic techniques. The local structure of sorbed U varied with loading, but the local structure of Sr did not vary with loading. Sorbed Sr exhibited specific adsorption as partially hydrated species whereas sorbed U exhibited specific adsorption as monomeric and dimeric U(VI)-carbonate complexes. Sorption proved site specific. These differences in site specificity and sorption mechanism may account forthe difficulties associated with predicting Sr and U loading and removal kinetics using MST.

Adsorption↗
PubMed2005

Probing the competition between secondary structures and local preferences in gas phase isolated peptide backbones.

Combining laser desorption with a supersonic expansion together with the selectivity of IR/UV double resonance spectroscopy makes it possible to isolate and characterise the gas phase of remarkable backbone conformations of short peptide chains mimicking protein segments. A systematic bottom-up approach involving a conformer-specific IR study of peptide sequences of increasing sizes has enabled us to map the spectral signatures of the intramolecular interactions, which shape the peptide backbone, in particular H-bonds. The precise data collected are directly comparable to the most sophisticated quantum chemistry calculations of these species and therefore constitute a stringent test for the theoretical methods used. One-residue chains reveal the local conformational preference of the backbone and its dependence upon the nature of the residue. The investigation of longer chains provides evidence for a competition between simple successions of local conformational preferences along the chain and more folded structures, in which a new H-bonding network, involving distant H-bonding sites along the backbone, takes place. From three residues, the issue of helical folding can also be addressed. The present review of the gas phase literature data emphasizes the observation of remarkable secondary structures of biology, including short segments of beta-strands, gamma- and beta-turns, combinations of turns, including a 3(10) helix. It also provides evidence for the flexibility of the peptide chains, i.e., a critical influence of rather minor interactions (like side-chain/backbone interactions) on the conformational stability. Finally, the paper will discuss future promising directions of the present approach.

Dipeptides↗
PubMed1996

The discriminability of local surface structure.

The ability of observers to discriminate depth and orientation differences between separated local regions on object surfaces was examined. The objects were defined by many optical sources of information simultaneously, including shading, texture, motion, and binocular disparity. Despite the full-cue nature of the displays, the observers' performance was relatively poor, with Weber fractions ranging from 10% to 40%. The Weber fractions were considerably lower for discriminations of surface-orientation differences than for similar discriminations of depth differences. The ability of observers to discriminate surface-orientation differences was approximately invariant over the separation of the regions in the projected image. In contrast, the ability to discriminate depth differences was highly influenced by the amount of image separation. This qualitative difference between the perception of depth intervals and surface-orientation differences suggests that knowledge of depths and orientations may be represented separately within the human visual system.

Depth Perception↗
PubMed1990

Carcinoembryonic antigen, its spatial structure and localization of antigenic determinants.

The effects of temperature and pH on the spatial structure of carcinoembryonic antigen (CEA) and its various derivatives are studied by circular dichroic spectroscopy and differential UV spectroscopy. The spatial organization of the protein portion of CEA and its derivatives as revealed by spectroscopic evidence is discussed with respect to the antigenic activity of the species studied. We conclude that the protein portion of CEA consists mainly of a beta-structural type. Using the various modifications of the sugar moiety and the protein portion, antigenic determinants of CEA are shown to possess the main conformational character of the molecule while participation of the sugars in the formation and orientation of the carbohydrate chains relative to the protein core of the antigen appear to be important.

Carbohydrate Conformation↗
PubMed2003

Annotation in three dimensions. PINTS: Patterns in Non-homologous Tertiary Structures.

The detection of local structural patterns in proteins (e.g. active sites) can provide insights into protein function in the absence of sequence or fold similarity. Methods to detect such similarities are key during structural annotation, for example with results from Structural Genomics initiatives. PINTS (Patterns in Non-homologous Tertiary Structures, http://pints.embl.de) performs database searches for such patterns and most importantly provides a measure of statistical significance for any similarity uncovered. To aid functional annotation of proteins, we allow comparisons of pre-defined patterns against databases of complete structures and of entire structures to databases of particular residues likely to be functionally important.

Binding Sites↗
PubMed1996

Chondroitinase ABC (pharmaceutical grade) for chemonucleolysis. Functional and structural evaluation after local application on intraspinal nerve structures and blood vessels.

STUDY DESIGN: The effects on nerve tissue and blood vessels of locally applied chondroitinase ABC were studied in two experimental models using chymopapain and the vehicle of chondroitinase ABC for controls. OBJECTIVES: To assess the effects of chondroitinase ABC on blood vessels and nerve tissue after local application. SUMMARY OF BACKGROUND DATA: Chondroitinase ABC has been suggested for chemonucleolysis because it has a high specificity for nucleus pulposus matrix, which could mean a high efficiency in dissolving disc tissue combined with a low risk of side effects on other tissues. METHODS: Chondroitinase ABC or controls were injected intrathecally in the pig, and nerve conduction velocity and histologic changes were assessed after 7 days. The same substances were injected into the hamster cheek pouch and studied for 60 minutes for microvascular effects. The vehicle for the enzyme was used as a negative control and chymopapain in a therapeutic concentration served as a positive control. RESULTS: In all series there was a slight intrathecal fibrotic reaction that was most pronounced after chymopapain injection. The effects on nerve conduction velocity and nerve morphology were similar between chondroitinase ABC and its vehicle. Chymopapain induced a significant reduction in nerve conduction velocity and pronounced histologic changes. In the cheek pouch, chymopapain induced a stand-still of blood flow at the injection site, and microhemorrhage and macromolecular leakage from the vessels at the border of the injection site. Only a slightly reduced blood flow was occasionally found after injection of chondroitinase ABC and controls. CONCLUSIONS: In agreement with the current literature, these observations indicate that chondroitinase ABC is safe regarding adverse effects on nerve tissue and blood vessels. The slight reduction in conduction velocity after intrathecal injection of chondroitinase ABC or its vehicle is most likely the result of surgical injury while releasing the nerve roots from the intrathecal fibrous adhesions. Such adhesions may be related to the laminectomy per se, and probably have no pathophysiologic significance.

Animals↗
PubMed2002

Role of framework sodium versus local framework structure in determining the hydrothermal stability of MCM-41 mesostructures.

Two mesostructured MCM-41 silicas that differ dramatically in hydrothermal stability have been examined by (29)Si MAS NMR spectroscopy and pair distribution function (PDF) analysis of synchrotron X-ray scattering data. The less stable mesostructure assembled from sodium silicate and the substantially more stable derivative made from fumed silica possess equivalent local framework wall structures, as judged by NMR and PDF methods. Approximately 80% of the SiO(4) tetrahedra are fully cross-linked as Q(4) (Si(OSi)(4)) units in both calcined samples. Additionally, the structural correlation distances for the two materials are nearly identical, having values of 1.62(1), approximately 2.60, and 3.09(1) A for the Si-O, O-O, and nearest neighbor Si-Si distances in the framework. Sodium ions in the framework play a crucial role in limiting the hydrothermal stability of the mesostructure. Residual sodium ( approximately 0.05-0.10% Na(2)O) is retained in the MCM-41 made from sodium silicate, even after two ion exchange reactions with ammonium ions in more than 300 - fold excess. The entrapped framework sodium ions catalyze the collapse of the mesopores upon exposure to 20% steam at 800 degrees C for 5 h. The sodium - free mesostructure assembled from fumed silica retains an open framework under the same hydrothermal conditions. The stability of the fumed silica derivative, however, is greatly compromised when doped with as little as 0.10% Na(2)O, thus confirming the deleterious effect of sodium on hydrothermal stability.

Journal Article↗
PubMed1992

Three-dimensional structure of proteolytic fragment 163-231 of bacterioopsin determined from nuclear magnetic resonance data in solution.

546 NOESY cross-peak volumes were measured in the two-dimensional NOESY spectrum of proteolytic fragment 163-231 of bacterioopsin in organic solution. These data and 42 detected hydrogen bonds were applied for determining the peptide spatial structure. The fold of the polypeptide chain was determined by local structure analysis, a distance geometry approach and systematic search for energetically allowed side-chain rotamers which are consistent with experimental NOESY cross-peak volumes. The effective rotational correlation time of 6 ns for the molecule was evaluated from optimization of the local structure to meet NOE data and from the dependence on mixing time of the NiH/Ci alpha H cross-peak volumes of the residues in alpha-helical conformation. The resulting structure has two well defined alpha-helical regions, 168-191 and 198-227, with root-mean-square deviation 44 pm and 69 pm, respectively, between the backbone atoms in 14 final energy refined conformations. The alpha-helices correspond to transmembrane segments F and G of bacteriorhodopsin. The segment F contains proline 186, which introduces a kink of about 25 degrees with a disruption of the hydrogen bond with the NH group of the following residue. The segments are connected by a flexible loop region 192-197. Torsion angles chi 1 are unequivocally defined for 62% of side chains in the alpha-helices but half of them differ from electron cryo-microscopy (ECM) model of bacteriorhodopsin, apparently because of the low resolution of ECM. Nevertheless, the F and G segments can be packed as in the ECM model and with side-chain conformations consistent with all NMR data in solution.

Amino Acid Sequence↗
PubMed2001

Gene structure, intracellular localization, and functional roles of sterol carrier protein-2.

Since its discovery three decades ago, sterol carrier protein-2 (SCP-2) has remained a fascinating protein whose physiological function in lipid metabolism remains an enigma. Its multiple proposed functions arise from its complex gene structure, post-translational processing, intracellular localization, and ligand specificity. The SCP-2 gene has two initiation sites coding for proteins that share a common 13 kDa SCP-2 C-terminus: (1) One site codes for 58 kDa SCP-x which is partially post-translationally cleaved to 13 kDa SCP-2 and a 45 kDa protein. (2) A second site codes for 15 kDa pro-SCP-2 which is completely post-translationally cleaved to 13 kDa SCP-2. Very little is yet known regarding how the relative proportions of the two transcripts are regulated. Although all three proteins contain a C-terminal SKL peroxisomal targeting sequence, it is unclear why all three proteins are not exclusively localized in peroxisomes. However, the recent demonstration that the SCP-2 N-terminal presequence in pro-SCP-2 dramatically modulated the intracellular targeting coded by the C-terminal peroxisomal targeting sequence may account for the observation that as much as half of total SCP-2 is localized outside the peroxisome. The tertiary and secondary structure of the 13 kDa SCP-2, but not that of 15 kDa pro-SCP-2 and 58 kDa SCP-x, are now resolved. Increasing evidence suggests that the 58 kDa SCP-x and 45 kDa proteins are peroxisomal 3-ketoacyl-CoA-thiolases involved in the oxidation of branched chain fatty acids. Since 15 kDa pro-SCP-2 is post-translationally completely cleaved to 13 kDa SCP-2, relatively little attention has been focused on this protein. Finally, although the 13 kDa SCP-2 is the most studied of these proteins, because it exhibits diversity of its ligand partners (fatty acids, fatty acyl CoAs, cholesterol, phospholipids), new potential physiological function(s) are still being proposed and questions regarding potential compensation by other proteins with overlapping specificity are only beginning to be resolved.

Acetyl-CoA C-Acetyltransferase↗
PubMed2004

Transmission electron microscopy and structural phase transitions in anion-deficient perovskite-based oxides.

This review deals with the structural transitions arising in anion-deficient Mn- and Cu-based perovskites when changing the anion content. Special attention is devoted to the local structure of such materials, studied with electron diffraction and high-resolution electron microscopy. Using examples of three-dimensional or layered manganites and cuprates, it is shown that oxidation/reduction reactions in such compounds are accompanied by the formation of domain structures. Fragmentation into domains can have intrinsic reasons, related to the loss of symmetry elements due to the ordering of anion vacancies, or because of the simultaneous realization of several different anion ordering patterns with close formation energies. When extra anions are inserted at low temperatures, limited diffusion ability also results in the appearance of domains of different structures arising from a local inhomogeneity in the anion distribution. The relevance of the investigation of the local structure for a correct structure interpretation is discussed.

Journal Article↗
PubMed1992

Conformation of parathyroid hormone: time-resolved fluorescence studies.

Conformational and environmental changes in the functionally significant amino-terminal region of human parathyroid hormone (hPTH), induced by solvent or by complexation with acidic lipid, have been investigated. Structural perturbations were monitored by their effect on the fluorescence decay kinetics of the single tryptophyl residue at position 23. Data for the intact hormone were compared with those for its 1-34 and 13-34 analogues. Deletion of the 35-84 sequence had no significant effect on the structure of hPTH in the region of Trp-23, nor was there any evidence for interaction of this region with the 1-12 sequence. On the basis of a comparison of the results of this study with structural information available from other spectroscopic techniques, we propose that the local structure in the region of Trp-23 of aqueous solutions of hPTH and hPTH 1-34 has helical character. This local structure was not stable in aqueous hPTH 13-34, but was present in hPTH and its analogues, both on complexation with acidic lipid and in helix-promoting solvents. The tryptophyl fluorescence of the lipid-bound peptides was characteristic of an aqueous environment. Triple-exponential fluorescence decay kinetics were observed for the tryptophyl residue of hPTH and its deletion analogues. This can be explained in terms of ground-state heterogeneity due to the presence of three C alpha-C beta rotamers of the tryptophanyl indole side chain. Assuming this model, we show that the calculated fractional concentrations of the decay time components correlate with the likely rotamer populations and with their expected dependence on the main-chain conformation.(ABSTRACT TRUNCATED AT 250 WORDS)

Circular Dichroism↗
PubMed1997

Human estrogen receptor beta-gene structure, chromosomal localization, and expression pattern.

The estrogen receptor (ER) is a ligand-activated transcription factor that mediates the effects of the steroid hormone 17 beta-estradiol, in both males and females. Since the isolation and cloning of ER, the consensus has been that only one such receptor exists. The finding of a second subtype of ER (ER beta) has caused considerable excitement amongst endocrinologists. In this article, we present data regarding the genomic structure and chromosomal localization of the human ER beta gene, demonstrating that two independent ER genes do exist in the human. Furthermore, we present data regarding the tissue distribution of human ER beta, showing that this receptor is expressed in multiple tissues. For instance, ER beta is found in developing spermatids of the testis, a finding of potential relevance for the ongoing debate on the effects of environmental estrogens on sperm counts. In addition, we find ER beta in ovarian granulosa cells, indicating that estrogens also participate in the regulation of follicular growth in the human.

Amino Acid Sequence↗