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At least 289 records · Page 16Linked to original sources
PubMed1991

Secondary structure analysis of the scrapie-associated protein PrP 27-30 in water by infrared spectroscopy.

A protease-resistant form of the protein PrP (PrP-res) accumulates in tissues of mammals infected with scrapie, Creutzfeldt-Jakob disease, and related transmissible neurodegenerative diseases. This abnormal form of PrP can aggregate into insoluble amyloid-like fibrils and plaques and has been identified as the major component of brain fractions enriched for scrapie infectivity. Using a recently developed technique in Fourier transform infrared spectroscopy which allows protein conformational analysis in aqueous media, we have studied the secondary structure of the proteinase K resistant core of PrP-res (PrP-res 27-30) as it exists in highly infectious fibril preparations. Second-derivative analysis of the infrared spectra has enabled us to quantitate the relative amounts of different secondary structures in the PrP-res aggregates. The analysis indicated that PrP-res 27-30 is predominantly composed of beta-sheet (47%), which is consistent with its amyloid-like properties. In addition, significant amounts of turn (31%) and alpha-helix (17%) were identified, indicating that amyloid-like fibrils need not be exclusively beta-sheet. The infrared-based secondary structure compositions were then used as constraints to improve the theoretical localization of the secondary structures within PrP-res 27-30.

Acetylglucosamine↗
PubMed1992

An antiplatelet aggregation principle and X-ray structural analysis of cis-khellactone diester from Peucedanum japonicum.

Three structurally related khellactone coumarins, 1-3, were isolated from the aerial parts of Peucedanum japonicum (Umbelliferae). Compound 2 was identified as a new coumarin (cis-3'-isovaleryl-4'-senecioylkhellactone) by spectral and chemical analysis. Conformation of the dihydropyrano ring of cis-3',4'-disenecioylkhellactone [3] was elucidated by X-ray crystallographic analysis. These three natural khellactone esters were subjected to the antiplatelet aggregation bioassay where cis-3',4'-diisovalerylkhellactone [1] showed significant activity (at 50 micrograms/ml).

Animals↗
PubMed2026

Molecular Cloning, Recombinant Expression, and In Silico Structural Analysis of Cu/Zn-Superoxide Dismutase from Trachyspermum ammi.

Superoxide dismutase (SOD) is an essential antioxidant metalloenzyme that is critical for the cellular defense against oxidative damage, as it scavenges superoxide radicals and maintains the redox status. Cytosolic Cu/Zn-SOD is particularly important in the regulation of oxidative stress among different isoforms in higher plants. While Cu/Zn-SODs from several plant species have been characterized, molecular information is limited for Trachyspermum ammi, a medicinally important member of a family Apiaceae with antioxidant potential.In the present study, an integrated molecular and in silico approach has been taken to clone and analyze a Cu/Zn type SOD gene from T. ammi to get insight into its structural and evolutionary characteristics. PCR amplification yielded an open reading frame of 456 bp encoding a protein of 152 amino acids. Sequence analysis showed that plant Cu/Zn-SODs, especially those from Daucus carota, were highly similar to one another (about 90-95%).Multiple sequence alignment confirmed the presence of conserved catalytic motifs and metal-binding histidine residues, both of which are crucial for enzymatic function. Physicochemical analysis predicted the protein to be stable, hydrophilic and compatible with cytosolic localization. The analysis of secondary structure indicated a predominance of β-strands, consistent with the conserved β-barrel architecture of plant Cu/Zn-SODs.The three-dimensional structure was built by homology modeling using a closely related plant Cu/Zn-SOD template with high sequence identity. Structural validation demonstrated an acceptable stereochemical quality with 86.3% residues in the favored region of Ramachandran plot, satisfactory ERRAT and Verify3D scores, and a low RMSD value of 0.104 Å on structural superimposition. Phylogenetic analysis placed the enzyme in the Apiaceae lineage, suggesting evolutionary conservation among related plant species. In conclusion, this study presents the first molecular and structural characterization of Cu/Zn-SOD from T. ammi and confirms the existence of a conserved structural framework typical of plant Cu/Zn-SODs. These results provide a basis for further studies concerning recombinant expression, enzymatic validation and potential relevance in antioxidant and plant stress biology.

Cloning, Molecular↗
PubMed1994

[Chemical structural analysis of steroids by NMR spectroscopy].

NMR is so convenient way to get stereochemical information of an organic compound that organic chemists have readily taken advantage of its spectra to investigate its stereochemistry by means of chemical shifts, coupling constants, nuclear Overhauser effect, etc. Especially in the steroid field, there have been reported many studies for determination of chemical structure and analysis of conformation. It is exemplified how NMR is in steroid chemistry.

Aldosterone↗
PubMed1991

Crystal structure analysis of the B-DNA dodecamer CGTGAATTCACG.

The crystal structure of the DNA dodecamer C-G-T-G-A-A-T-T-C-A-C-G has been determined at a resolution of 2.5 A, with a final R factor of 15.8% for 1475 nonzero reflections measured at 0 degrees C. The structure is isomorphous with that of the Drew dodecamer, with the space group P2(1)2(1)2(1) and cell dimensions of a = 24.94 A, b = 40.78 A, and c = 66.13 A. The asymmetric unit contains all 12 base pairs of the B-DNA double helix and 36 water molecules. The structure of C-G-T-G-A-A-T-T-C-A-C-G is very similar to that of C-G-C-G-A-A-T-T-C-G-C-G, with no major alterations in helix parameters. Water peaks in the refined structure appear to represent a selection of peaks that were observed in the Drew dodecamer. The minor-groove spine of hydration at 2.5 A is fragmentary, but as Narendra et al. (1991) [Biochemistry (following paper in this issue)] have observed, lowering the temperature leads to a more complete representation of the spine.

Base Sequence↗
PubMed1993

Nucleic acid structure analysis: a users guide to a collection of new analysis programs.

Common nomenclature describing the geometry of nucleic acid structures was established at a 1988 EMBO Workshop on DNA Curvature and Bending (Diekmann, S. (1988) J. Mol. Biol. 208, 787-791; Diekmann, S. (1989) The EMBO Journal 8, 1-4; Sarma, R.H. (1988) J. Biomol. Structure & Dynamics 6, 391-395; Dickerson, R.E. (1989) J. Biomol. Structure & Dynamics 6, 627-634; Dickerson, R.E. et al. (1989) Nuc. Acids Res. 17, 1979-1803). We have subsequently developed and incorporated sophisticated mathematics in a computer program to calculate the parameters described by the guidelines. The program calculates all the local parameters relating complementary bases and neighboring base and base pairs in both Cartesian and helical coordinate frames. In addition, the main mathematical property requested by the EMBO guidelines--that the magnitude of the parameters be independent of strand or direction of measurement--is accomplished without the use of a midway coordinate frame for the rotational parameters. The mathematics preserve the physical intuition used in defining the parameters; in particular, the rotational parameters are true rotations based on a simple physical model (rotation at constant angular velocity for a unit amount of time), not Euler angles or angles between vectors and planes as is the case with other approaches. As a result, the mathematical equations are symmetric with the property that a 5 degrees tilt is the same as a 5 degrees roll or a 5 degrees twist, except that the rotations take place about different axes. In other approaches, a 5 degrees tilt can mean a different amount of net rotation from a 5 degrees roll or a 5 degrees twist. In addition, a great deal of flexibility is built into the program so that the user has control over the analysis, including the input format, the coordinate frame used for the base pairing relationship, the point about which the rotations are performed, and which geometric relationships are analyzed. While there is a great deal of flexibility, the program is easy to use. Interactive queries and user accessible files make the options in the program very convenient to tailor to individual needs. In addition, there is also a program that calculates bond lengths, valence angles, and torsion angles along the nucleic acid backbone, and within the sugar and base rings. Another program 'learns' the identities of the bond lengths, valence angles, and torsion angles that the user would like to determine.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗
PubMed1992

Three-dimensional structure of acylphosphatase. Refinement and structure analysis.

We report here the complete determination of the solution structure of acylphosphatase, a small enzyme that catalyses the hydrolysis of organic acylphosphates, as determined by distance geometry methods based on nuclear magnetic resonance information. A non-standard strategy for the distance geometry calculations was used and is described here some detail. The five best structures were then refined by restrained energy minimization and molecular dynamics in order to explore the conformational space consistent with the experimental data. We address the question of whether the solution structure of acylphosphatase follows the general principles of protein structure, i.e. those learned from analysing crystal structures. Static and dynamic features are discussed in detail. An uncommon beta-alpha-beta motif, so far found only in procarboxypeptidase B and in an RNA-binding protein, is present in acylphosphatase.

Acid Anhydride Hydrolases↗
PubMed1991

Three-dimensional structural analysis of the group B polysaccharide of Neisseria meningitidis 6275 by two-dimensional NMR: the polysaccharide is suggested to exist in helical conformations in solution.

The solution conformations of the group B polysaccharide of Neisseria meningitidis were analyzed by DQF-COSY and pure absorption 2D NOE NMR with three mixing times. The pyranose ring of the sialic acid residue was found to be in the 2C5 conformation. The DQF-COSY analysis indicated that the orientations of H6 and H7 and of H7 and H8 are both gauche. In order to overcome the difficulties in analyzing the NOE data due to the two sets of proton overlaps, molecular modeling of alpha-2,8-linked sialic acid oligomers was carried out to investigate possible conformers, and theoretical NOE calculations were performed by using CORMA (complete relaxation matrix analysis). Our analysis suggests that the polysaccharide adopts helical structures for which the phi (defined by O6-C2-O8-C8) and psi (C2-O8-C8-C7) angles are in the following ranges: phi -60 to 0 degrees, psi 115-175 degrees or phi 90-120 degrees, psi 55-175 degrees. The weak affinity of anti-B antibodies for smaller alpha-2,8-linked oligosaccharides may be due to the fact that such oligomers are more flexible and may not form an ordered structure as the poly(sialic acid) does.

Antigens, Bacterial↗
PubMed1985

Human T-cell lymphotropic virus type III: immunologic characterization and primary structure analysis of the major internal protein, p24.

The major internal structural protein of human T-cell lymphotropic virus type III (HTLV-III), a virus etiologically implicated in acquired immunodeficiency syndrome (AIDS), was purified to homogeneity. This 24,000-molecular-weight protein (p24) was shown to lack immunologic cross-reacting antigenic determinants shared by other known retroviruses, including HTLV-I and HTLV-II, with the exception of equine infectious anemia virus (EIAV). A broadly reactive competition immunoassay was developed in which antiserum to EIAV was used to precipitate 125I-labeled HTLV-III p24. Although the major structural proteins of HTLV-III and EIAV competed in this assay, other type B, C, and D retroviral proteins lacked detectable reactivity. Thus, HTLV-III is more related to EIAV than to any other retroviruses. That the HTLV-III isolate is very distinct from HTLV-I and HTLV-II was further confirmed by the amino acid compositions of the major internal antigens of all three isolates. Moreover, comparison of the amino-terminal amino acid sequence of HTLV-III p24 with analogous sequences for HTLV-I and HTLV-II p24 showed that these proteins do not share significant sequence homology. In an attempt to evaluate immune response in individuals exposed to HTLV-III, sera from AIDS and lymphadenopathy syndrome patients as well as from clinically normal blood donor controls were tested for antibodies to HTLV-III p24. The results showed that sera from 93% of lymphadenopathy syndrome patients and 73% of AIDS patients exhibited high-titered antibodies to HTLV-III p24. In contrast, none of the normal control sera showed detectable reactivity to HTLV-III p24.

Acquired Immunodeficiency Syndrome↗
PubMed1984

Human alpha 1-proteinase inhibitor. Crystal structure analysis of two crystal modifications, molecular model and preliminary analysis of the implications for function.

Two closely related crystal structures of alpha 1-proteinase inhibitor modified at the reactive site peptide bond Met358--Ser359 have been analysed. The crystal structure has been obtained from diffraction data at 3 A resolution, with phases originally from isomorphous replacement. The electron density map was substantially improved by cyclic averaging of the electron densities of the two crystal forms and allowed the chain to be traced in terms of the known chemical amino acid sequence. Energy restrained crystallographic refinement was initiated and resulted in conventional R-values of 0.251 for the tetragonal crystal form (6 to 3 A resolution) and 0.247 for the hexagonal crystal form (6 to 3.2 A resolution). The polypeptide chain is almost completely arranged in well-defined secondary structural elements: three beta-sheets and eight alpha-helices. The helices are preferentially formed by the first 150 residues. They are in proximity underneath sheet A. The chain ends Met358 and Ser359 of the nicked species are arranged in strands on opposite ends of the molecule indicating a major structural rearrangement upon modification of the intact inhibitor. It is suggested that the Met358 strand is in a different conformation removed from sheet A and approaches Ser359 in the intact inhibitor species. Glu342, which is exchanged by a lysine in the Z-variant is in a strategic position for such a rearrangement. The three carbohydrate chains of alpha 1-proteinase inhibitor have partly defined electron density close to their attachment sites at asparagine residues. The anti-thrombin and ovalbumin amino acid sequences can be accommodated in the alpha 1 inhibitor molecular structure. The intron-exon junctions of the ovalbumin and the alpha 1-proteinase inhibitor gene are all in surface loops of the mature protein.

Amino Acid Sequence↗
PubMed1993

Structural analysis of the purine repressor, an Escherichia coli DNA-binding protein.

The purine repressor protein, PurR, is a member of the lac repressor, LacI, family of Escherichia coli DNA-binding proteins that bind DNA via a highly conserved N-terminal helix-turn-helix motif. Additionally, the members of this family display strong sequence homologies between their larger C-terminal effector binding/oligomerization domains. Analysis of the PurR primary and secondary structures reveals that its C-terminal corepressor binding domain is highly homologous to another group of E. coli-binding proteins, the periplasmic binding proteins, particularly to the ribose-binding protein (RBP). The high resolution x-ray structure of RBP allows this protein to serve as a template with which to model the predicted secondary structure of the corepressor binding domain of PurR. Similarly, the N-terminal DNA binding domain of PurR can be modeled using the NMR-determined structure of the corresponding region (residues 1-56) from LacI as a template. Combining the two, results in a description of the likely secondary structure topology of PurR and implicates residues important for corepressor binding and dimerization. CD spectroscopic studies on PurR, its corepressor binding domain and RBP result in secondary structure estimates nearly identical with those obtained by sequence analyses, thereby providing further corroborating physical evidence for this topological assignment.

Amino Acid Sequence↗
PubMed1995

Chromatin structure analysis based on a hierarchic texture model.

The quantification of chromatin structures is an important part of nuclear grading of malignant and premalignant lesions. In order to achieve high accuracy, computerized image analysis systems have been applied in this process. Chromatin texture analysis of cell nuclei requires a suitable texture model. A hierarchic model seemed to be most compatible for this purpose. It assumes that texture consists of homogeneous regions (textons). Based on this model, two approaches to texture segmentation and feature extraction were investigated using sections of cervical tissue. We examined the reproducibility of the measurement under changing optical conditions. The coefficients of variations of the texture features ranged from 2.1% to 16.9%. The features were tested for their discriminating capability in a pilot study including 30 cases of cervical dysplasia and carcinoma. The overall classification accuracy reached 65%. This study presents an automated technique for texture analysis that is similar to human perception.

Carcinoma in Situ↗
PubMed1993

Cloning of the cDNA encoding human xanthine dehydrogenase (oxidase): structural analysis of the protein and chromosomal location of the gene.

The primary structure of human xanthine dehydrogenase (hXDH) was determined by cloning and sequence analysis of the cDNAs encoding the enzyme. The nucleotide (nt) sequence has an open reading frame of 3999 nt encoding a protein of 1333 amino acids (aa) with a calculated M(r) of 146,604. The deduced aa sequence of hXDH is homologous to the previously reported rat XDH (rXDH) and Drosophila melanogaster XDH sequences with identities of 90.2 and 52.0%, respectively. The aa residues involved in both the reversible and the irreversible conversion from the dehydrogenase type to the oxidase type of rXDH are completely conserved between the rat and the human enzymes. This implies that the molecular mechanisms of the conversion of hXDH from dehydrogenase to oxidase are common to those of the well-characterized rXDH. Five sequence variations were detected in the isolated cDNA clones. Spot blot hybridization using flow-sorted human chromosome revealed that the hXDH-encoding gene (hXDH) was located on chromosome 2.

Amino Acid Sequence↗
PubMed1992

Design and synthesis of novel 6,7-imidazotetrahydroquinoline inhibitors of thymidylate synthase using iterative protein crystal structure analysis.

Antifolate inhibitors of thymidylate synthase (TS) have primarily been based on the structure of folic acid. This paper describes the identification and development of novel 6,7-imidazotetrahydroquinoline TS inhibitors by iterative ligand design, synthesis, and crystallographic analysis of protein-inhibitor complexes. Beginning with a high-resolution crystal structure of E. coli TS (TS, EC 2.1.1.45), an imidazotetrahydroquinoline inhibitor was designed de novo to occupy the folate binding pocket. Structural modifications of the initial compound 1h (Ki approximately 5 microM human/E. coli TS) were then made on the basis of feedback from additional cocrystal structures and activity data. An amino group in the 2-position of the imidazole was found to increase the potency of the series by 1-2 orders of magnitude. Other substitutions on the imidazole ring (1-CH3, 2-CH3, 2-NHCH3, 2-SCH3) generally led to weaker inhibition. Additional improvements in activity were obtained by modification of the substituents on the tetrahydroquinoline nitrogen, bringing the Ki of three of the compounds below 15 nM against the human TS enzyme. The compounds were tested for cytotoxicity and were shown to inhibit the growth of three tumor cell lines in vitro.

Adenocarcinoma↗
PubMed1984

[Structural analysis of the correlations of genetic markers with physiological indices].

The structure of phenotypic and genetic correlations between ABO blood groups, MN, Rh(D), haptoglobins and concentration indications of the total cholesterol, of sugar curve, the levels of systolic and diastolic arterial pressure was studied in twins by the method for cluster analysis (121 monozygotic twin pairs, 120 dizygotic like sex twin pairs and 107 pairs of unlike sex twins). No correlation between the systems analysed and their indications on the phenotypic level was observed. By analysis of genetic correlations, the existence of correlations between the Rh system and arterial pressure, and the total cholesterol concentration, as well as between the ABO system and sugar concentration indexes, was established. Possible genetic causes for correlations obtained are being discussed.

Adult↗
PubMed1984

Structural analysis of heparin by methylation and g.l.c.-m.s.: preliminary results.

Heparin is a complex mixture of polysaccharides differing in biological activity and structure, and attempts to relate this activity to structure have suffered, owing to a lack of sufficiently sensitive and specific analytical methods. Application of methylation analysis to determination of the structure of heparin is described. Carboxyl-reduced heparin was converted into its pyridinium salt, this was dissolved in Me2SO, and free OH and NH groups were methylated with dimethylsulfinyl anion. Sulfate groups were removed by solvolysis, and after dialysis, the polymer was acetylated and depolymerized by acetolysis. The resulting monosaccharides were converted into alditol acetates, which were separated by capillary, gas-liquid chromatography, and identified by both electron impact and chemical ionization mass spectrometry. Seventeen different monosaccharides were identified in the hydrolyzate. All of the expected internal hexosaminyl and glycosyluronic residues were identified. Although several sugars were identified as nonreducing termini, only a hexosamine 6-sulfate was identified as a reducing-terminus sugar. The results indicate that methylation analysis of heparins and other complex, sulfated glycosaminoglycans is feasible.

Chemical Phenomena↗
PubMed1994

Structural analysis of antibody specificity. Detailed comparison of five Fab'-steroid complexes.

Structures of the Fab' fragment of the anti-progesterone antibody DB3 in complex with five cross-reactive steroids (aetiocholanolone, 5 beta-androstane-3,17-dione, 5 alpha-pregnane-20-one-3 beta-ol-hemisuccinate, progesterone-11 alpha-ol-hemisuccinate and progesterone) have been determined by X-ray crystallography to a maximum resolution of 2.7 A. These different steroids compete with progesterone binding with affinities in the nanomolar range despite substantial differences in their three-dimensional structures. Comparison of the unliganded DB3 Fab' and these five steroid-Fab' complexes reveals that all the steroid ligands bind to an "open" conformation of the Fab' as defined by the orientation of the indole side-chain of TrpH100, whereas in the unliganded or "closed" form the binding site is occluded by TrpH100. Small but significant conformational changes take place in the antibody to maximize the physical and chemical complementarity with each ligand. The various cross-reactive ligands are accommodated in the binding site in two distinct orientations. We term these binding modes syn and anti, as they are defined by the orientation of the steroid beta face relative to TrpH50. In all cases, the steroid D ring is inserted into a hydrophobic cavity formed mainly by TrpH50, TyrH97, TrpH100 and PheH100b; a hydrogen bond interaction with AsnH35 to the keto group at position C17 or C20 orients the steroid in the pocket. The AsnH35 hydrogen bond and the interaction with TrpH50 account for the restricted heavy chain response to immunization with progesterone-like steroids derivatized at the 11 alpha position. Cross-reactivity of the antibody with different steroids is explained by alternative binding pockets for the A ring, which generates different ligand orientations in the binding site. This study suggests which factors are most likely to contribute to the observed antibody specificity, such as linker position and the paucity of functional groups on the immunogenic hapten.

Amino Acid Sequence↗
PubMed1993

Bisamides from Aglaia species: structure analysis and potential to reverse drug resistance with cultured cells.

The structure of pyramidatine [1], a new bisamide alkaloid from leaves of Aglaia pyramidata, was determined through extensive nmr studies, including homonuclear COSY, NOESY, APT, HETCOR, and selective INEPT techniques. Revision of the 13C-nmr assignment of piriferine [2], an alkaloid previously isolated from A. pirifera, was achieved by examination of several 2D nmr spectra (homonuclear COSY, NOESY, and HETCOR) and confirmed by selective INEPT nmr experiments. Evaluation of the cytotoxic potential of the two alkaloids, along with two other bisamides from Aglaia odorata, odorine [3] and 5'-epi-odorine [4], was carried out in eleven human cancer cell lines. None of these bisamides showed significant cytotoxicity. Nevertheless, piriferine [2], odorine [3], and 5'-epi-odorine [4] were found to inhibit the growth of the vinblastine-resistant KB cells by enhancing the anticancer activity of vinblastine.

Alkaloids↗