共查询到20条相似文献,搜索用时 15 毫秒
1.
SA Moore BF Anderson CR Groom M Haridas EN Baker 《Canadian Metallurgical Quarterly》1997,274(2):222-236
The three-dimensional structure of diferric bovine lactoferrin (bLf) has been determined by X-ray crystallography in order to investigate the factors that influence iron binding and release by transferrins. The structure was solved by molecular replacement, using the coordinates of diferric human lactoferrin (hLf) as a search model, and was refined with data to 2.8 A resolution by simulated annealing (X-PLOR) and restrained least squares (TNT). The final model comprises 5310 protein atoms (residues 5 to 689), 124 carbohydrate atoms (from ten monosaccharide units, in three glycan chains), 2 Fe3+, 2 CO32- and 50 water molecules. This model gives an R-factor of 0.232 for 21440 reflections in the resolution range 30.0 to 2.8 A. The folding of the bLf molecule is essentially the same as that of hLf, but bLf differs in the extent of closure of the two domains of each lobe, and in the relative orientations of the two lobes. Differences in domain closure are attributed to amino acid changes in the interface, and differences in lobe orientations to slightly altered packing of two hydrophobic patches between the lobes. Changed interdomain interactions may explain the lesser iron affinity of bLf, compared with hLf, and two lysine residues behind the N-lobe iron site of bLf offer new insights into the "dilysine trigger" mechanism proposed for iron release by transferrins. The bLf structure is also notable for several well-defined oligosaccharide units which demonstrate the structural factors that stabilise carbohydrate structure. One glycan chain, attached to Asn545, appears to contribute to interdomain interactions and may modulate iron release from the C-lobe. 相似文献
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G Tsiotis M Psylinakis B Woplensinger A Lustig A Engel D Ghanotakis 《Canadian Metallurgical Quarterly》1999,259(1-2):320-324
The photosystem II (PSII) reaction center (RC) complex was isolated from spinach and characterized by gel electrophoresis, gel filtration and analytical ultracentrifugation. The purified complex contained the PsbA, PsbD, PsbE, PsbF and PsbI subunits. Gel filtration and analytical ultracentrifugation indicated the presence of a homogeneous complex. The mass of the RC complexes was found to be 107 kDa by analytical ultracentrifugation and 132 kDa by scanning transmission electron microscopy (STEM). The mass obtained showed the isolated complex to exist as a monomer and only one cytochrome b559 (cyt b559) to be associated with the RC complex. Digital images of negatively stained RC complexes were recorded by STEM and analyzed by single-particle averaging. The complex was 9 nm long and 5 nm wide, and exhibited a pronounced quasi-twofold symmetry. This supports the symmetric organization of the PSII complex, with the PsbA and the PsbD proteins in the center and symmetrically arranged PsbB and PsbC proteins at the periphery of the monomeric complex. 相似文献
3.
K Mayanagi T Ishikawa C Toyoshima Y Inoue K Nakazato 《Canadian Metallurgical Quarterly》1998,123(3):211-224
Risk factors for central venous catheter (CVC)-related bacteraemia among infants admitted to a neonatal intensive care unit (NICU) were analysed and the impact of surveillance and continuing education on the incidence of this complication investigated. Among patients admitted to a NICU, CVC-related bacteraemia increased from 1/15 (7%) in 1987 to 11/26 (42%) in 1988 (P = 0.01). Coagulase-negative staphylococci isolated from bacteraemia patients showed clonal diversity by plasmid and chromosomal fingerprinting. A review of CVC care procedures suggested breaches in aseptic techniques. Catheter-care technique was revised to ensure maximal aseptic precautions, including the use of sterile gloves, gown and drapes. The new policy was promoted by a continuing education programme and regular feed-back of CVC-related bacteraemia incidence to NICU staff. In the four-year follow-up period, the attack-rate of CVC-related bacteraemia decreased to 18/156 (12%) patients [relative risk (RR): 0.27, 95% confidence interval (CI); 0.15-0.51; P < 0.001 vs the previous period]. By using the Cox's model proportional hazards, very low birthweight and the period before use of strict aseptic CVC care were found to be predictors of increased risk of catheter-related bacteraemia after adjustment for duration of catheterization. These data provide further evidence that strict aseptic precautions during the maintenance and utilization of CVC can contribute to lower the risk of catheter infection in critically ill neonates. Regular feedback of surveillance data was associated with a progressive decrease in incidence of infection, suggesting that it improved staff compliance with aseptic precautions. 相似文献
4.
A Tuuttila E Morgunova U Bergmann Y Lindqvist K Maskos C Fernandez-Catalan W Bode K Tryggvason G Schneider 《Canadian Metallurgical Quarterly》1998,284(4):1133-1140
The three-dimensional structure of human tissue inhibitor of metalloproteinases-2 (TIMP-2) was determined by X-ray crystallography to 2.1 A resolution. The structure of the inhibitor consists of two domains. The N-terminal domain (residues 1-110) is folded into a beta-barrel, similar to the oligonucleotide/oligosaccharide binding fold otherwise found in certain DNA-binding proteins. The C-terminal domain (residues 111-194) contains a parallel stranded beta-hairpin plus a beta-loop-beta motif. Comparison of the structure of uncomplexed human TIMP-2 with that of bovine TIMP-2 bound to the catalytic domain of human MMP-14 suggests an internal rotation between the two domains of approximately 13 degrees upon binding to the protease. Furthermore, local conformational differences in the two structures that might be induced by formation of the protease-inhibitor complex have been found. The most prominent of these involves residues 27-40 of the A-B beta-hairpin loop. Structure-based alignment of amino acid sequences of representatives of the TIMP family maps the sequence differences mainly to loop regions, and some of these differences are proposed to be responsible for the particular properties of the various TIMP species. 相似文献
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The pigment content of isolated reaction centers of photosystem II was modified using an exchange protocol similar to that used for purple bacterial reaction centers. With this method, which is based on incubation of reaction centers at elevated temperature with an excess of chemically modified pigments, it was possible to incorporate [3-acetyl]-chlorophyll a and [Zn]-chlorophyll a into photosystem II reaction centers. Pigment exchange has been verified by absorption, circular dichroism and fluorescence spectroscopy, and quantitated by HPLC analysis of pigment extracts. 相似文献
8.
A Mattevi G Obmolova KH Kalk WJ van Berkel WG Hol 《Canadian Metallurgical Quarterly》1993,230(4):1200-1215
The structure of Pseudomonas fluorescens lipoamide dehydrogenase, a dimeric flavoenzyme with a molecular mass of 106,000 daltons, was solved by the molecular replacement method and refined to an R-factor of 19.4% at 2.8 A resolution. The root-mean-square difference from ideal values for bonds and angles is 0.019 A and 3.8 degrees, respectively. The structure is closely related to that of the same flavoprotein from Azotobacter vinelandii. The root-mean-square difference for 932 C alpha atoms is 0.64 A, with 84% sequence identity. The residues in the active site are identical, while 89% of the interface residues are the same in the two enzymes. A few structural variations provide the basis for the differences in thermostability and redox properties between the two homologous proteins. Particularly, in the A. vinelandii molecule a threonine to alanine (T452A) mutation leaves a buried carbonyl oxygen, located at the subunit interface and in proximity of the flavin ring, unpaired to any H-bond donor, probably providing an explanation for the lower stability of the A. vinelandii enzyme with respect to the P. fluorescens enzyme. Six surface loops, which previously could not be accurately positioned in the A. vinelandii structure, are well defined in P. fluorescens lipoamide dehydrogenase. On the basis of the P. fluorescens structure, the six loops could be correctly defined also in the A. vinelandii enzyme. This is an unusual case where similar refinement methodologies applied to two crystal forms of closely related proteins led to electron density maps of substantially different quality. The correct definition of these surface residues is likely to be an essential step for revealing the structural basis of the interactions between lipoamide dehydrogenase and the other members of the pyruvate dehydrogenase multienzyme complex. 相似文献
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A number of extracellular proteins contain cryptic inhibitors of angiogenesis. Endostatin is a 20 kDa C-terminal proteolytic fragment of collagen XVIII that potently inhibits endothelial cell proliferation and angiogenesis. Therapy of experimental cancer with endostatin leads to tumour dormancy and does not induce resistance. We have expressed recombinant mouse endostatin and determined its crystal structure at 1.5 A resolution. The structure reveals a compact fold distantly related to the C-type lectin carbohydrate recognition domain and the hyaluronan-binding Link module. The high affinity of endostatin for heparin is explained by the presence of an extensive basic patch formed by 11 arginine residues. Endostatin may inhibit angiogenesis by binding to the heparan sulphate proteoglycans involved in growth factor signalling. 相似文献
11.
K Luger AW M?der RK Richmond DF Sargent TJ Richmond 《Canadian Metallurgical Quarterly》1997,389(6648):251-260
The X-ray crystal structure of the nucleosome core particle of chromatin shows in atomic detail how the histone protein octamer is assembled and how 146 base pairs of DNA are organized into a superhelix around it. Both histone/histone and histone/DNA interactions depend on the histone fold domains and additional, well ordered structure elements extending from this motif. Histone amino-terminal tails pass over and between the gyres of the DNA superhelix to contact neighbouring particles. The lack of uniformity between multiple histone/DNA-binding sites causes the DNA to deviate from ideal superhelix geometry. 相似文献
12.
A Telfer S Dhami SM Bishop D Phillips J Barber 《Canadian Metallurgical Quarterly》1994,33(48):14469-14474
By measuring time-resolved luminescence emission at 1270 nm, we have detected singlet oxygen formation by illuminated, reaction centers of photosystem II isolated from Pisum sativum, which is in agreement with earlier work (Macpherson, A. N., Telfer, A., Barber, J., & Truscott, T. G. (1993) Biochim. Biophys. Acta 1143, 301-309). In this paper we show that the yield of singlet oxygen is significantly increased if the number of beta-carotene molecules bound per isolated complex is reduced from two to one. We conclude, therefore, that beta-carotene can act as an effective quencher of singlet oxygen in the photosystem II reaction center. This conclusion is supported by the finding that the rate of light-induced irreversible bleaching of chlorins in the reaction center is increased with decreasing beta-carotene levels. The results demonstrate the direct intermediacy of singlet oxygen in causing photooxidative damage within a biological environment and are discussed, specifically, in terms of the role of beta-carotene in protecting photosystem II against photoinhibition. 相似文献
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The refined structure of dimeric diphtheria toxin (DT) at 2.0 A resolution, based on 37,727 unique reflections (F > 1 sigma (F)), yields a final R factor of 19.5% with a model obeying standard geometry. The refined model consists of 523 amino acid residues, 1 molecule of the bound dinucleotide inhibitor adenylyl 3'-5' uridine 3' monophosphate (ApUp), and 405 well-ordered water molecules. The 2.0-A refined model reveals that the binding motif for ApUp includes residues in the catalytic and receptor-binding domains and is different from the Rossmann dinucleotide-binding fold. ApUp is bound in part by a long loop (residues 34-52) that crosses the active site. Several residues in the active site were previously identified as NAD-binding residues. Glu 148, previously identified as playing a catalytic role in ADP-ribosylation of elongation factor 2 by DT, is about 5 A from uracil in ApUp. The trigger for insertion of the transmembrane domain of DT into the endosomal membrane at low pH may involve 3 intradomain and 4 interdomain salt bridges that will be weakened at low pH by protonation of their acidic residues. The refined model also reveals that each molecule in dimeric DT has an "open" structure unlike most globular proteins, which we call an open monomer. Two open monomers interact by "domain swapping" to form a compact, globular dimeric DT structure. The possibility that the open monomer resembles a membrane insertion intermediate is discussed. 相似文献
14.
M Gajhede DJ Schuller A Henriksen AT Smith TL Poulos 《Canadian Metallurgical Quarterly》1997,4(12):1032-1038
The crystal structure of horseradish peroxidase isozyme C (HRPC) has been solved to 2.15 A resolution. An important feature unique to the class III peroxidases is a long insertion, 34 residues in HRPC, between helices F and G. This region, which defines part of the substrate access channel, is not present in the core conserved fold typical of peroxidases from classes I and II. Comparison of HRPC and peanut peroxidase (PNP), the only other class III (higher plant) peroxidase for which an X-ray structure has been completed, reveals that the structure in this region is highly variable even within class III. For peroxidases of the HRPC type, characterized by a larger FG insertion (seven residues relative to PNP) and a shorter F' helix, we have identified the key residue involved in direct interactions with aromatic donor molecules. HRPC is unique in having a ring of three peripheral Phe residues, 142, 68 and 179. These guard the entrance to the exposed haem edge. We predict that this aromatic region is important for the ability of HRPC to bind aromatic substrates. 相似文献
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Squalene cyclases catalyze a cationic cyclization cascade, which is homologous to a key step in cholesterol biosynthesis. The structure of the enzyme from Alicyclobacillus acidocaldarius has been determined in a new crystal form at 2.0 A resolution (1 A=0.1 nm) and refined to an R-factor of 15.3 % (Rfree=18.7 %). The structure indicates how the initial protonation and the final deprotonation of squalene occur and how the transient carbocations are stabilized. The pathways of the flexible educt squalene from the membrane interior to the active center cavity and of the rigid fused-ring product hopene in the reverse direction are discussed. The enzyme contains eight so-called QW-sequence repeats that fortify the alpha/alpha-barrels by an intricate interaction network. They are unique to the known triterpene cyclases and are presumed to shield these enzymes against the released enthalpy of the highly exergonic catalyzed reaction. The enzyme is a monotopic membrane protein, the membrane-binding interactions of which are described and compared with those of two prostaglandin-H2 synthase isoenzymes, the only other structurally characterized proteins of this type. In the crystals the membrane-binding regions face each other, suggesting a micelle-type detergent structure between them. 相似文献
16.
The DNA-containing capsid of canine parvovirus (CPV) is analyzed following atomic refinement at 2.9 A resolution. The capsid contains 60 copies of the capsid protein related by icosahedral symmetry. The atomic model has been extended from the first residue (Gly37) of the unrefined 3.25 A structure towards the N terminus. The electron density shows that approximately 87% of the capsid proteins have N termini on the inside of the capsid, but for approximately 13%, the polypeptide starts on the outside and runs through one of the pores surrounding each 5-fold axis, explaining apparently conflicting antigenic data. Analysis of potential hydrogen bonds reveals approximately 50% more secondary structure than previously apparent. Most of the additional secondary structure are in the 71 and 221 residue-long loop insertions between beta-strands E and F and G and H, forming subunit-bridging sheets that likely add specificity to assembly interactions. Structural analysis of the extensive subunit interactions around the 3-fold axes shows that assembly is a multistep process with loops intertwining following initial contact. Estimated free energies of association suggest that the formation of 3 and 5-fold contacts likely takes precedence over 2-fold interactions. Energies for initial association into trimers or pentamers would be similar, but the intertwining of loops about the 3-fold axis adds an additional large activation barrier to dissociation. Analysis of the surfaces of the assembled capsid shows a surprising lack of basic amino acids that might have been expected to interact with the negatively charged phosphoribose backbone of the DNA. Instead, uncharged polar and van der Waal's interactions predominate in the packaging of single-stranded DNA into the capsid. 相似文献
17.
ML Groot van Mourik F C Eijckelhoff van Stokkum IHM JP Dekker van Grondelle R 《Canadian Metallurgical Quarterly》1997,94(9):4389-4394
In photosystem II of green plants the key photosynthetic reaction consists of the transfer of an electron from the primary donor called P680 to a nearby pheophytin molecule. We analyzed the temperature dependence of this reaction by subpicosecond transient absorption spectroscopy over the temperature range 20-240 K using isolated photosystem II reaction centers from spinach. After excitation in the red edge of the Qy absorption band, the decay of the excited state can conveniently be described by two kinetic components that both accelerate with temperature. This temperature behavior differs remarkably from that observed in purple bacterial reaction centers. We attribute the first component, which accelerates from 2.6 ps at 20 K to 0.4 ps at 240 K, to charge separation after direct excitation of P680, and explain its temperature dependence by an intermediate that lies in energy above the singlet-excited P680 and that possibly has charge-transfer character. The second component accelerates from 120 ps at 20 K to 18 ps at 240 K and is attributed to charge separation after direct excitation of the "trap" state near-degenerate with P680 and subsequent slow energy transfer from this trap state to P680. We suggest that the slow energy transfer from the trap state to P680 plays an important role in the kinetics of radical pair formation at room temperature. 相似文献
18.
ES Cedergren-Zeppezauer NC Goonesekere MD Rozycki JC Myslik Z Dauter U Lindberg CE Schutt 《Canadian Metallurgical Quarterly》1994,240(5):459-475
Profilin regulates the behavior of the eukaryotic microfilament system through its interaction with non-filamentous actin. It also binds several ligands, including poly(L-proline) and the membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). Bovine profilin crystals (space group C2; a = 69.15 A, b = 34.59 A, c = 52.49 A; alpha = gamma = 90 degrees, beta = 92.56 degrees) were grown from a mixture of poly(ethylene glycol) 400 and ammonium sulfate. X-ray diffraction data were collected on an imaging plate scanner at the DORIS storage ring (DESY, Hamburg), and were phased by molecular replacement, using a search model derived from the 2.55 A structure of profilin complexed to beta-actin. The refined model of bovine profilin has a crystallographic R-factor of 16.5% in the resolution range 6.0 to 2.0 A and includes 128 water molecules, several of which form hydrogen bonds to stabilize unconventional turns. The structure of free bovine profilin is similar to that of bovine profilin complexed to beta-actin, and C alpha atoms from the two structures superimpose with an r.m.s. deviation of 1.25 A. This value is reduced to 0.51 A by omitting Ala1 and the N-terminal acetyl group, which lie at a profilin-actin interface in crystals of the complex. These residues display a strained conformation in crystalline profilin-actin but may allow the formation of a hydrogen bond between the N-acetyl carbonyl group of profilin and the phenol hydroxyl group of Tyr188 in actin. Several other actin-binding residues of profilin show different side-chain rotomer conformations in the two structures. The polypeptide fold of bovine profilin is generally similar to those observed by NMR for profilin from other sources, although the N terminus of Acanthamoeba profilin isoform I lies in a distorted helix and the C-terminal helix is less tilted with respect to the strands in the central beta-pleated sheet than is observed in bovine profilin. The majority of the aromatic residues in profilin are exposed to solvent and lie in either of two hydrophobic patches, neither of which takes part in an interface with actin. One of these patches is required for binding poly(L-proline) and contains an aromatic cluster comprising the highly conserved residues Trp3, Tyr6, Trp31 and Tyr139. In forming this cluster, Trp31 adopts a sterically strained rotamer conformation.(ABSTRACT TRUNCATED AT 400 WORDS) 相似文献
19.
KZ Pan YJ Lin KJ Zhou ZJ Fu MH Chen DR Huang DH Huang 《Canadian Metallurgical Quarterly》1993,36(9):1069-1081
The crystallographic refinement of trichosanthin has been performed at 2.6 A resolution. The crystal and molecular structure of trichosanthin is described in detail in this paper. On summarizing the regularity of the amino acid sequences of eight kinds of ribosome inactivating proteins and combining with the crystal and molecular structure of trichosanthin, fifteen most conservative amino acid residues are analyzed. It is found that four most conservative polar amino acid residues Gln156, Glu160, Arg163 and Glu189 gather on the molecular surface on the boundary of the large and small domains, thus forming the active center of the protein molecule. 相似文献
20.
S Strobl K Maskos M Betz G Wiegand R Huber FX Gomis-Rüth R Glockshuber 《Canadian Metallurgical Quarterly》1998,278(3):617-628
The three-dimensional structure of the alpha-amylase from Tenebrio molitor larvae (TMA) has been determined by molecular replacement techniques using diffraction data of a crystal of space group P212121 (a=51.24 A; b=93.46 A; c=96.95 A). The structure has been refined to a crystallographic R-factor of 17.7% for 58,219 independent reflections in the 7.0 to 1.64 A resolution range, with root-mean-square deviations of 0.008 A for bond lengths and 1.482 degrees for bond angles. The final model comprises all 471 residues of TMA, 261 water molecules, one calcium cation and one chloride anion. The electron density confirms that the N-terminal glutamine residue has undergone a post-transitional modification resulting in a stable 5-oxo-proline residue. The X-ray structure of TMA provides the first three-dimensional model of an insect alpha-amylase. The monomeric enzyme exhibits an elongated shape approximately 75 Ax46 Ax40 A and consists of three distinct domains, in line with models for alpha-amylases from microbial, plant and mammalian origin. However, the structure of TMA reflects in the substrate and inhibitor binding region a remarkable difference from mammalian alpha-amylases: the lack of a highly flexible, glycine-rich loop, which has been proposed to be involved in a "trap-release" mechanism of substrate hydrolysis by mammalian alpha-amylases. The structural differences between alpha-amylases of various origins might explain the specificity of inhibitors directed exclusively against insect alpha-amylases. 相似文献