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At least 1,621 records · Page 90Linked to original sources
PubMed1999

Determination of a protein structure by iodination: the structure of iodinated acetylxylan esterase.

Enzymatic and non-enzymatic iodination of the amino acid tyrosine is a well known phenomenon. The iodination technique has been widely used for labeling proteins. Using high-resolution X-ray crystallographic techniques, the chemical and three-dimensional structures of iodotyrosines formed by non-enzymatic incorporation of I atoms into tyrosine residues of a crystalline protein are described. Acetylxylan esterase (AXE II; 207 amino-acid residues) from Penicillium purpurogenum has substrate specificities towards acetate esters of D-xylopyranose residues in xylan and belongs to a new class of alpha/beta hydrolases. The crystals of the enzyme are highly ordered, tightly packed and diffract to better than sub-angström resolution at 85 K. The iodination technique has been utilized to prepare an isomorphous derivative of the AXE II crystal. The structure of the enzyme determined at 1.10 A resolution exclusively by normal and anomalous scattering from I atoms, along with the structure of the iodinated complex at 1.80 A resolution, demonstrate the formation of covalent bonds between I atoms and C atoms at ortho positions to the hydroxyl groups of two tyrosyl moieties, yielding iodotyrosines.

Acetylesterase↗
PubMed2005

Two crystal structures of dihydrofolate reductase-thymidylate synthase from Cryptosporidium hominis reveal protein-ligand interactions including a structural basis for observed antifolate resistance.

Cryptosporidium hominis is a protozoan parasite that causes acute gastrointestinal illness. There are no effective therapies for cryptosporidiosis, highlighting the need for new drug-lead discovery. An analysis of the protein-ligand interactions in two crystal structures of dihydrofolate reductase-thymidylate synthase (DHFR-TS) from C. hominis, determined at 2.8 and 2.87 A resolution, reveals that the interactions of residues Ile29, Thr58 and Cys113 in the active site of C. hominis DHFR provide a possible structural basis for the observed antifolate resistance. A comparison with the structure of human DHFR reveals active-site differences that may be exploited for the design of species-selective inhibitors.

Animals↗
PubMed2004

Experimentally biased model structure of the Hsc70/auxilin complex: substrate transfer and interdomain structural change.

A model structure of the Hsc70/auxilin complex has been constructed to gain insight into interprotein substrate transfer and ATP hydrolysis induced conformational changes in the multidomain Hsc70 structure. The Hsc70/auxilin system, which is a member of the Hsp70/Hsp40 chaperone system family, uncoats clathrin-coated vesicles in an ATP hydrolysis-driven process. Incorporating previous results from NMR and mutant binding studies, the auxilin J-domain was docked into the Hsc70 ATPase domain lower cleft using rigid backbone/flexible side chain molecular dynamics, and the Hsc70 substrate binding domain was docked by a similar procedure. For comparison, J-domain and substrate binding domain docking sites were obtained by the rigid-body docking programs DOT and ZDOCK, filtered and ranked by the program ClusPro, and relaxed using the same rigid backbone/flexible side chain dynamics. The substrate binding domain sites were assessed in terms of conserved surface complementarity and feasibility in the context of substrate transfer, both for auxilin and another Hsp40 protein, Hsc20. This assessment favors placement of the substrate binding domain near D152 on the ATPase domain surface adjacent to the J-domain invariant HPD segment, with the Hsc70 interdomain linker in the lower cleft. Examining Hsc70 interdomain energetics, we propose that long-range electrostatic interactions, perhaps due to a difference in the pKa values of bound ATP and ADP, could play a major role in the structural change induced by ATP hydrolysis. Interdomain electrostatic interactions also appear to play a role in stimulation of ATPase activity due to J-domain binding and substrate binding by Hsc70.

Adenosine Triphosphate↗
PubMed2001

Genomic-scale comparison of sequence- and structure-based methods of function prediction: does structure provide additional insight?

A function annotation method using the sequence-to-structure-to-function paradigm is applied to the identification of all disulfide oxidoreductases in the Saccharomyces cerevisiae genome. The method identifies 27 sequences as potential disulfide oxidoreductases. All previously known thioredoxins, glutaredoxins, and disulfide isomerases are correctly identified. Three of the 27 predictions are probable false-positives. Three novel predictions, which subsequently have been experimentally validated, are presented. Two additional novel predictions suggest a disulfide oxidoreductase regulatory mechanism for two subunits (OST3 and OST6) of the yeast oligosaccharyltransferase complex. Based on homology, this prediction can be extended to a potential tumor suppressor gene, N33, in humans, whose biochemical function was not previously known. Attempts to obtain a folded, active N33 construct to test the prediction were unsuccessful. The results show that structure prediction coupled with biochemically relevant structural motifs is a powerful method for the function annotation of genome sequences and can provide more detailed, robust predictions than function prediction methods that rely on sequence comparison alone.

Algorithms↗
PubMed1982

Conformation and structure of acidic dipeptides. Crystal structure of glycyl-L-aspartic acid dihydrate.

The crystal structure of the acidic dipeptide glycyl-L-aspartic acid dihydrate, Gly-L-Asp X 2H2O, C6H10N2O5 X 2H2O, has been determined by means of three-dimensional counter X-ray data. The dipeptide crystallizes in space group P212121 of the orthorhombic system with four formula units in a cell of dimensions of a = 9.611(4), b = 9.775(6), and c = 10.845(4)A. The structure was solved by direct methods and refined by least-squares methods to a final value of the weighted R-factor of 0.035 using all 1191 observed intensity data. The dipeptide occurs in the crystal as a zwitterion, with the glycyl N-terminus protonated and the aspartyl main chain carboxyl group deprotonated. The conformation about the peptide bond is fully trans extended, with an omega angle of -175.9 degrees. While there is extensive intermolecular hydrogen bonding in the crystals, there is no intramolecular hydrogen bonding either here or in the solid state structures of related dipeptides.

Dipeptides↗
PubMed1987

Examination of the structural requirements for proliferation of peroxisomes and mitochondria in mouse liver by hypolipidemic agents, with special emphasis on structural analogues of 2-ethylhexanoic acid.

We have found here that there are clear structural requirements for peroxisome proliferation (monitored as increases in carnitine acetyltransferase activity, cyanide-insensitive palmitoyl-CoA oxidation, catalase and increases in the protein designated PPA 80) in mouse liver. From the investigation of ten structural analogues of 2-ethylhexanoic acid, it could be concluded that the most effective proliferators all have an ethyl group as the substituent on carbon 2 of the main chain, which consists of six carbons. The further observation from this group of compounds that a charged group is required for effective proliferation leads us to speculate that such a group is involved in the molecular mechanism as well. Many, but not all, of the effective peroxisome proliferators in a second group of compounds contain a phenoxy group, often with a substituted alpha carbon. Interestingly, the 2,4-dichlorophenoxyacetic and 2,4,5-trichlorophenoxyacetic acids are both effective peroxisome proliferators, but the closely related p-chlorophenoxyacetic acid is inactive in this respect, indicating that the chlorine atom at position 2 must be essential to the process in these cases. The results presented here also indicate that the structural requirements for proliferation of mitochondria are similar to those for proliferation of peroxisomes. Certainly, the most effective peroxisome proliferators also cause large increases in 'mitochondrial' protein and cytochrome oxidase activity, i.e. there is an obvious qualitative correlation.

Animals↗
PubMed1993

Glucose-6-phosphate dehydrogenase. Structure-function relationships and the Pichia jadinii enzyme structure.

The primary structure of glucose-6-phosphate dehydrogenase from the yeast Pichia jadinii (formerly Candida utilis) has been determined. It consists of a 495-residue, N-terminally acetylated protein chain. The structure shows extensive differences from those of the corresponding mammalian, fruit fly, and bacterial enzymes (52-68% residue non-identities), but also from that of another yeast, Saccharomyces cerevisiae (38%). A eubacterial type and a yeast type of glucose-6-phosphate dehydrogenase are discerned, in addition to the known mammalian type. They are distinguished from each other, from the mammalian type, and the insect enzyme, on the basis of both specific residues and pattern differences. The distribution of residues conserved in all forms locates short segments in which identities are closely grouped. Approximately 50% of these segments correspond to predicted turns and appear to mark the principal folds characteristic of the enzyme's tertiary structure. A region in the N-terminal part of the protein chain has characteristics suggestive of a coenzyme-binding site, while, in the middle third, another functionally important segment may be related to glucose-6-phosphate binding and catalysis.

Amino Acid Sequence↗
PubMed1996

Solution 1H-NMR structure of the heme cavity in the low-affinity state for the allosteric monomeric cyano-met hemoglobins from Chironomus thummi thummi. Comparison to the crystal structure.

Solution 1H-NMR studies of the heme cavity were performed for the cyanomet complexes of monomeric hemoglobins III and IV from the insect Chironomus thummi thummi, each of which exhibit marked Bohr effects. The low pH 5, paramagnetic (S = 1/2) derivatives were selected for study because the large dipolar shifts provide improved resolution over diamagnetic forms and allow distinction between the two isomeric heme orientations [Peyton, D. H., La Mar, G. N. & Gersonde, K. (1988) Biochim. Biophys. Acta 954, 82-94]. The crystal structure for the low-pH form of the hemoglobin III derivative, moreover, has been reported and showed that the functionally implicated distal His58 side chain adopts alternative orientation, either in or out of the pocket [Steigemann, W. & Weber, E. (1979) J. Mol. Biol. 127, 309-338]. All heme pocket residues for the low-pH forms of the two hemoglobins were located, at least in part, and positioned in the heme cavity on the basis of nuclear Overhauser effects to the heme and each other, dipolar shifts, and paramagnetic-induced relaxation. The resulting structure yielded the orientation of the major axis of the paramagnetic susceptibility tensor. The heme pocket structure of the cyanomet hemoglobins III and IV were found to be indistinguishable, with both exhibiting a distal His58 oriented solely into the heme cavity and in contact with the ligand, and with two residues, Phe100 and Phe38, exhibiting small but significant displacements in solution relative to hemoglobin III in the crystal.

Allosteric Site↗
PubMed1997

Elucidation of the structure of the core region and the complete structure of the R-type lipopolysaccharide of Erwinia carotovora FERM P-7576.

An R-type lipopolysaccharide (LPS) from Erwinia carotovora strain FERM P-7576 was studied after strong alkaline degradation and mild acid hydrolysis. The resulting products were analyzed by fast-atom bombardment mass spectrometry, one- and two-dimensional 1H and 13C NMR spectroscopy, dephosphorylation and methylation analysis. The following structure was proposed for the core region of the LPS: [formula in text] where Hep is L-glycero-D-manno-heptose and Kdo is 3-deoxy-D-manno-octulosonic acid. Some LPS species lack the beta-D-Galp residue or the beta-D-Galp-(1-->7)-alpha-Hepp disaccharide. With the known structures of lipid A [Fukuoka, S., Kamishima, H., Nagawa, Y., Nakanishi, H., Ishikawa, K., Niwa, Y., Tamiya, E. & Karube, I. (1992) Arch. Microbiol. 157, 311-318] and the core moiety, the complete LPS structure was established and confirmed by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry of the native and O-deacylated LPS.

Carbohydrate Conformation↗
PubMed2002

Pharmacophore definition and three-dimensional quantitative structure-activity relationship study on structurally diverse prostacyclin receptor agonists.

Prostacyclin is an endogenous mediator that shows potent platelet inhibitory activity and powerful relaxation of peripheral resistance vessels. Prostacyclin receptor agonists are valuable drugs in the treatment of various vascular diseases spanning primary pulmonary hypertension to Raynaud's syndrome. Although agonists from various structural classes were synthesized, a common pharmacophore was never defined. Therefore, an attempt was made to integrate the different agonists into a single model. A dataset of structurally diverse prostacyclin receptor agonists was tested for its affinity to the human platelet prostacyclin receptor. The dataset included prostanoid and nonprostanoid ligands comprising iloprost, cicaprost, and BMY45778. Extensive conformational analyses were performed for both classes of compounds because of the absence of rigid templates. The search and superimposition procedure yielded a pharmacophore that aligns the essential carboxylate group of the agonists as well as demonstrates that different functional groups in prostanoid and nonprostanoid agonists can be arranged in a uniform conformation. A three-dimensional quantitative structure-activity relationship study was performed using the programs GRID and GOLPE. This analysis yielded a cross-validated correlation coefficient of 0.77. With this model, it is possible to predict the affinity of untested compounds.

Binding Sites↗
PubMed2003

Crystal structure of a putative methyltransferase from Mycobacterium tuberculosis: misannotation of a genome clarified by protein structural analysis.

Bioinformatic analyses of whole genome sequences highlight the problem of identifying the biochemical and cellular functions of many gene products that are at present uncharacterized. The open reading frame Rv3853 from Mycobacterium tuberculosis has been annotated as menG and assumed to encode an S-adenosylmethionine (SAM)-dependent methyltransferase that catalyzes the final step in menaquinone biosynthesis. The Rv3853 gene product has been expressed, refolded, purified, and crystallized in the context of a structural genomics program. Its crystal structure has been determined by isomorphous replacement and refined at 1.9 A resolution to an R factor of 19.0% and R(free) of 22.0%. The structure strongly suggests that this protein is not a SAM-dependent methyltransferase and that the gene has been misannotated in this and other genomes that contain homologs. The protein forms a tightly associated, disk-like trimer. The monomer fold is unlike that of any known SAM-dependent methyltransferase, most closely resembling the phosphohistidine domains of several phosphotransfer systems. Attempts to bind cofactor and substrate molecules have been unsuccessful, but two adventitiously bound small-molecule ligands, modeled as tartrate and glyoxalate, are present on each monomer. These may point to biologically relevant binding sites but do not suggest a function. In silico screening indicates a range of ligands that could occupy these and other sites. The nature of these ligands, coupled with the location of binding sites on the trimer, suggests that proteins of the Rv3853 family, which are distributed throughout microbial and plant species, may be part of a larger assembly binding to nucleic acids or proteins.

Amino Acid Sequence↗
PubMed2005

Structural characterization and assembly of the distal tail structure of the temperate lactococcal bacteriophage TP901-1.

The tail structures of bacteriophages infecting gram-positive bacteria are largely unexplored, although the phage tail mediates the initial interaction with the host cell. The temperate Lactococcus lactis phage TP901-1 of the Siphoviridae family has a long noncontractile tail with a distal baseplate. In the present study, we investigated the distal tail structures and tail assembly of phage TP901-1 by introducing nonsense mutations into the late transcribed genes dit (orf46), tal(TP901-1) (orf47), bppU (orf48), bppL (orf49), and orf50. Transmission electron microscopy examination of mutant and wild-type TP901-1 phages showed that the baseplate consisted of two different disks and that a central tail fiber is protruding below the baseplate. Evaluation of the mutant tail morphologies with protein profiles and Western blots revealed that the upper and lower baseplate disks consist of the proteins BppU and BppL, respectively. Likewise, Dit and Tal(TP901-1) were shown to be structural tail proteins essential for tail formation, and Tal(TP901-1) was furthermore identified as the tail fiber protein by immunogold labeling experiments. Determination of infection efficiencies of the mutant phages showed that the baseplate is fundamental for host infection and the lower disk protein, BppL, is suggested to interact with the host receptor. In contrast, ORF50 was found to be nonessential for tail assembly and host infection. A model for TP901-1 tail assembly, in which the function of eight specific proteins is considered, is presented.

Genes, Viral↗
PubMed2002

Aura virus structure suggests that the T=4 organization is a fundamental property of viral structural proteins.

Aura and Sindbis viruses are closely related alphaviruses. Unlike other alphaviruses, Aura virus efficiently encapsidates both genomic RNA (11.8 kb) and subgenomic RNA (4.2 kb) to form virus particles. Previous studies on negatively stained Aura virus particles predicted that there were two major size classes with potential T=3 and T=4 capsid structures. We have used cryoelectron microscopy and three-dimensional image reconstruction techniques to examine the native morphology of different classes of Aura virus particles produced in BHK cells. Purified particles separated into two components in a sucrose gradient. Reconstructions of particles in the top and bottom components were computed to resolutions of 17 and 21 A, respectively, and compared with reconstructions of Sindbis virus and Ross River virus particles. Aura virus particles of both top and bottom components have similar, T=4 structures that resemble those of other alphaviruses. The morphology of Aura virus glycoprotein spikes closely resembles that of Sindbis virus spikes and is detectably different from that of Ross River virus spikes. Thus, some aspects of the surface structure of members of the Sindbis virus lineage have been conserved, but other aspects have diverged from the Semliki Forest/Ross River virus lineage.

Alphavirus↗
PubMed1999

Structural characterization of the 16-kDa allergen, RA17, in rice seeds. Prediction of the secondary structure and identification of intramolecular disulfide bridges.

The 16-kDa rice allergen, RA17, belonging to the alpha-amylase/trypsin inhibitor family was isolated from rice seed and structurally characterized by identifying cystine-containing peptides and predicting the secondary structure and hydrophobic regions. Eight peptides, which constitute three sets of cystine-containing peptides, were purified by HPLC from a thermolytic digest of RA17 and identified by their amino acid sequence and composition, indicating five intramolecular disulfide bridges: Cys34-Cys94, Cys26-(Cys50 or Cys51)-Cys110 and Cys12-(Cys62 or Cys64)-Cys122. Analyses of the CD spectrum and the Chou-Fasman prediction suggested that RA17 had some helical- and sheet-structure regions. Based on these experimental and predicted data, RA17 is proposed to be a globular molecule with a small hydrophobic core having folding restricted by five intramolecular disulfide bridges.

Allergens↗
PubMed1985

Structure-induction versus structure-toxicity relationships for polychlorinated biphenyls and related aromatic hydrocarbons.

A comparison of the structure-induction (involving rat and mouse Ah receptor binding) and structure-toxicity (in vivo guinea pig toxicity) relationships suggests that two receptors with structurally distinct binding properties may be involved. This is supported by demonstration of potentiated toxicity through a mechanism believed to involve the Ah receptor as a site of loss with respect to toxicity. Theoretical and working models are proposed for these separate receptors to aid in the search for other relevant binding proteins. The findings suggest that polychlorinated biphenyls that are relatively low in toxicity may have modulating properties on the action of highly toxic compounds with which they are normally found in the environment.

Animals↗
PubMed2003

Determinants of enzymatic specificity in the Cys-Met-metabolism PLP-dependent enzymes family: crystal structure of cystathionine gamma-lyase from yeast and intrafamiliar structure comparison.

The crystal structure of cystathionine gamma-lyase (CGL) from yeast has been solved by molecular replacement at a resolution of 2.6 A. The molecule consists of 393 amino acid residues and one PLP moiety and is arranged in the crystal as a tetramer with D2 symmetry as in other related enzymes of the Cys-Met-metabolism PLP-dependent family like cystathionine beta-lyase (CBL). A structure comparison with other family members revealed surprising insights into the tuning of enzymatic specificity between the different family members. CGLs from yeast or human are virtually identical at their active sites to cystathionine gamma-synthase (CGS) from E. coli. Both CGLs and bacterial CGSs exhibit gamma-synthase and gamma-lyase activities depending on their position in the metabolic pathway and the available substrates. This group of enzymes has a glutamate (E333 in yeast CGL) which binds to the distal group of cystathionine (CTT) or the amino group of cysteine. Plant CGSs use homoserine phosphate instead of O-succinyl-homoserine as one substrate. This is reflected by a partially different active site structure in plant CGSs. In CGL and CBL the pseudosymmetric substrate must dock at the active site in different orientations, with S in gamma-position (CBL) or in delta-position (CGL). The conserved glutamate steers the substrate as seen in other CGLs. In CBLs this position is occupied by either tyrosine or hydrophobic residues directing binding of CTT such that S is in the in gamma-position. In methionine gamma-lyase a hydrophic patch operates as recognition site for the methyl group of the methionine substrate.

Binding Sites↗
PubMed2006

Crystal structure of the Bowman-Birk Inhibitor from Vigna unguiculata seeds in complex with beta-trypsin at 1.55 A resolution and its structural properties in association with proteinases.

The structure of the Bowman-Birk inhibitor from Vigna unguiculata seeds (BTCI) in complex with beta-trypsin was solved and refined at 1.55 A to a crystallographic R(factor) of 0.154 and R(free) of 0.169, and represents the highest resolution for a Bowman-Birk inhibitor structure to date. The BTCI-trypsin interface is stabilized by hydrophobic contacts and hydrogen bonds, involving two waters and a polyethylene glycol molecule. The conformational rigidity of the reactive loop is characteristic of the specificity against trypsin, while hydrophobicity and conformational mobility of the antichymotryptic subdomain confer the self-association tendency, indicated by atomic force microscopy, of BTCI in complex and free form. When BTCI is in binary complexes, no significant differences in inhibition constants for producing a ternary complex with trypsin and chymotrypsin were detected. These results indicate that binary complexes present no conformational change in their reactive site for both enzymes confirming that these sites are structurally independent. The free chymotrypsin observed in the atomic force microscopy assays, when the ternary complex is obtained from BTCI-trypsin binary complex and chymotrypsin, could be related more to the self-association tendency between chymotrypsin molecules and the flexibility of the reactive site for this enzyme than to binding-related conformational changes.

Amino Acid Sequence↗
PubMed2003

Quantitative structure-activity relationship models for prediction of estrogen receptor binding affinity of structurally diverse chemicals.

The demonstrated ability of a variety of structurally diverse chemicals to bind to the estrogen receptor has raised the concern that chemicals in the environment may be causing adverse effects through interference with nuclear receptor pathways. Many structure-activity relationship models have been developed to predict chemical binding to the estrogen receptor as an indication of potential estrogenicity. Models based on either two-dimensional or three-dimensional molecular descriptions that have been used to predict potential for binding to the estrogen receptor are the subject of the current review. The utility of such approaches to predict binding potential of diverse chemical structures in large chemical inventories, with potential application in a tiered risk assessment scheme, is discussed.

Animals↗