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1.
J B Finley V R Atigadda F Duarte J J Zhao W J Brouillette G M Air M Luo 《Journal of molecular biology》1999,293(5):1107-1119
The active site of type A or B influenza virus neuraminidase is composed of 11 conserved residues that directly interact with the substrate, sialic acid. An aromatic benzene ring has been used to replace the pyranose of sialic acid in our design of novel neuraminidase inhibitors. A bis(hydroxymethyl)pyrrolidinone ring was constructed in place of the N-acetyl group on the sialic acid. The hydroxymethyl groups replace two active site water molecules, which resulted in the high affinity of the nanomolar inhibitors. However, these inhibitors have greater potency for type A influenza virus than for type B influenza virus. To resolve the differences, we determined the X-ray crystal structure of three benzoic acid substituted inhibitors bound to the active site of B/Lee/40 neuraminidase. The investigation of a hydrophobic aliphatic group and a hydrophilic guanidino group on the aromatic inhibitors shows changes in the interaction with the active site residue Glu275. The results provide an explanation for the difference in efficacy of these inhibitors against types A and B viruses, even though the 11 active site residues of the neuraminidase are conserved. 相似文献
2.
Sequence and structure alignment of paramyxovirus hemagglutinin-neuraminidase with influenza virus neuraminidase. 总被引:22,自引:7,他引:15
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A model is proposed for the three-dimensional structure of the paramyxovirus hemagglutinin-neuraminidase (HN) protein. The model is broadly similar to the structure of the influenza virus neuraminidase and is based on the identification of invariant amino acids among HN sequences which have counterparts in the enzyme-active center of influenza virus neuraminidase. The influenza virus enzyme-active site is constructed from strain-invariant functional and framework residues, but in this model of HN, it is primarily the functional residues, i.e., those that make direct contact with the substrate sialic acid, which have identical counterparts in neuraminidase. The framework residues of the active site are different in HN and in neuraminidase and appear to be less strictly conserved within HN sequences than within neuraminidase sequences. 相似文献
3.
Three-dimensional structure of the neuraminidase of influenza virus A/Tokyo/3/67 at 2.2 A resolution 总被引:18,自引:0,他引:18
An atomic model of the tetrameric surface glycoprotein neuraminidase of influenza virus A/Tokyo/3/67 has been built and refined based on X-ray diffraction data at 2.2 A resolution. The crystallographic residual is 0.21 for data between 6 and 2.2 A resolution and the r.m.s. deviations from ideal geometry are 0.02 A for bond lengths and 3.9 degrees for bond angles. The model includes amino acid residues 83 to 469, four oligosaccharide structures N-linked at asparagine residues 86, 146, 200 and 234, a single putative Ca2+ ion site, and 85 water molecules. One of the oligosaccharides participates in a novel crystal contact. The folding pattern is a beta-sheet propeller as described earlier and details of the intramolecular interactions between the six beta-sheets are presented. Strain-invariant residues are clustered around the propeller axis on the upper surface of the molecule where they line the wall of a cavity into which sialic has been observed to bind. Strain-variable residues implicated in binding to antibodies surround this site. 相似文献
4.
Nandi T 《Bioinformation》2008,2(6):240-244
Human infection with avian influenza H5N1 is an emerging infectious disease characterized by respiratory symptoms and a high fatality rate. Hemagglutinin and neuraminidase are the two surface proteins responsible for infection by influenza virus. Till date, neuraminidase has been the major target for antiviral drugs. In the present study we chose hemagglutinin protein as it mediates the binding of the virus to target cells through sialic acid residues on the host cell-surface. Hemagglutinin of H5 avian influenza (PDB ID: 1JSN) was used as the receptor protein. Ligands were generated by structure-based de novo approach and virtual screening of ZINC database. A total of 11,104 conformers were generated and docked into the receptor binding site using 'High Throughput Virtual Screening'. We proposed potential lead molecules against the receptor binding site of hemagglutinin based on the results obtained from in silico docking and hydrogen bond interaction between the ligand and the 1JSN protein molecule. We found sialic acid derivative 1 to be the lead molecules amongst the ligands generated by structure based de novo approach. However the molecules obtained from ZINC database were showing better docking scores as well as conserved hydrogen bond interactions. Thus we proposed ZINC00487720 and ZINC00046810 as potential lead molecules that could be used as an inhibitor to the receptor binding site of hemagglutinin. They could now be studied in vivo to validate the in silico results. 相似文献
5.
Three-dimensional structure of the complex of 4-guanidino-Neu5Ac2en and influenza virus neuraminidase. 总被引:6,自引:1,他引:5
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J. N. Varghese V. C. Epa P. M. Colman 《Protein science : a publication of the Protein Society》1995,4(6):1081-1087
The three-dimensional X-ray structure of a complex of the potent neuraminidase inhibitor 4-guanidino-Neu5Ac2en and influenza virus neuraminidase (Subtype N9) has been obtained utilizing diffraction data to 1.8 A resolution. The interactions of the inhibitor, solvent water molecules, and the active site residues have been accurately determined. Six water molecules bound in the native structure have been displaced by the inhibitor, and the active site residues show no significant conformational changes on binding. Sialic acid, the natural substrate, binds in a half-chair conformation that is isosteric to the inhibitor. The conformation of the inhibitor in the active site of the X-ray structure concurs with that obtained by theoretical calculations and validates the structure-based design of the inhibitor. Comparison of known high-resolution structures of neuraminidase subtypes N2, N9, and B shows good structural conservation of the active site protein atoms, but the location of the water molecules in the respective active sites is less conserved. In particular, the environment of the 4-guanidino group of the inhibitor is strongly conserved and is the basis for the antiviral action of the inhibitor across all presently known influenza strains. Differences in the solvent structure in the active site may be related to variation in the affinities of inhibitors to different subtypes of neuraminidase. 相似文献
6.
Influenza Hemagglutinin and Neuraminidase Membrane Glycoproteins 总被引:2,自引:0,他引:2
Considerable progress has been made toward understanding the structural basis of the interaction of the two major surface glycoproteins of influenza A virus with their common ligand/substrate: carbohydrate chains terminating in sialic acid. The specificity of virus attachment to target cells is mediated by hemagglutinin, which acquires characteristic changes in its receptor-binding site to switch its host from avian species to humans. Anti-influenza drugs mimic the natural sialic acid substrate of the virus neuraminidase enzyme but utilize the much tighter binding of the drugs for efficacy. Resistance to one of the two main antiviral drugs is differentially acquired by the two distinct subsets of neuraminidase as a consequence of structural differences in the enzyme active site between the two phylogenetic groups. 相似文献
7.
Site-directed mutation of the active site of influenza neuraminidase and implications for the catalytic mechanism 总被引:6,自引:0,他引:6
Different isolates of influenza virus show a high degree of amino acid sequence variation in their surface glycoproteins. Conserved residues located in the substrate-binding pocket of the influenza virus neuraminidase are therefore likely to be involved in substrate binding or enzyme catalysis. In order to study the structure and function of the active site of this protein, a full-length cDNA clone of the neuraminidase gene from influenza A/Tokyo/3/67 was subcloned into aN M13 vector and amino acid substitutions were made in selected residues by using the oligonucleotide mismatch technique. The mutant neuraminidase genes were expressed from a recombinant SV40 vector, and the proteins were analyzed for synthesis, transport to the cell surface, and proper three-dimensional folding by internal and surface immunofluorescence. The mutant neuraminidase proteins were then assayed to determine the effect of the amino acid substitution on enzyme activity. Twelve of the 14 mutant proteins were correctly folded and were transported to the cell surface in a manner identical with that of the wild type. Two of these have full enzyme activity, but seven mutants, despite correct three-dimensional structure, have completely lost neuraminidase activity. Two mutants were active at low pH. The properties of the mutant enzymes suggest a possible mechanism of neuraminidase action. 相似文献
8.
Carolina Q. Sacramento Andressa Marttorelli Natalia Fintelman-Rodrigues Caroline S. de Freitas Gabrielle R. de Melo Marco E. N. Rocha Carlos R. Kaiser Katia F. Rodrigues Gisela L. da Costa Cristiane M. Alves Osvaldo Santos-Filho Jussara P. Barbosa Thiago Moreno L. Souza 《PloS one》2015,10(10)
The influenza virus causes acute respiratory infections, leading to high morbidity and mortality in groups of patients at higher risk. Antiviral drugs represent the first line of defense against influenza, both for seasonal infections and pandemic outbreaks. Two main classes of drugs against influenza are in clinical use: M2-channel blockers and neuraminidase inhibitors. Nevertheless, because influenza strains that are resistant to these antivirals have been described, the search for novel compounds with different mechanisms of action is necessary. Here, we investigated the anti-influenza activity of a fungi-derived natural product, aureonitol. This compound inhibited influenza A and B virus replication. This compound was more effective against influenza A(H3N2), with an EC50 of 100 nM. Aureonitol cytoxicity was also very low, with a CC50 value of 1426 μM. Aureonitol inhibited influenza hemagglutination and, consequently, significantly impaired virus adsorption. Molecular modeling studies revealed that aureonitol docked in the sialic acid binding site of hemagglutinin, forming hydrogen bonds with highly conserved residues. Altogether, our results indicate that the chemical structure of aureonitol is promising for future anti-influenza drug design. 相似文献
9.
Refined atomic structures of N9 subtype influenza virus neuraminidase and escape mutants 总被引:8,自引:0,他引:8
W R Tulip J N Varghese A T Baker A van Donkelaar W G Laver R G Webster P M Colman 《Journal of molecular biology》1991,221(2):487-497
The crystal structure of the N9 subtype neuraminidase of influenza virus was refined by simulated annealing and conventional techniques to an R-factor of 0.172 for data in the resolution range 6.0 to 2.2 A. The r.m.s. deviation from ideal values of bond lengths is 0.014 A. The structure is similar to that of N2 subtype neuraminidase both in secondary structure elements and in their connections. The three-dimensional structures of several escape mutants of neuraminidase, selected with antineuraminidase monoclonal antibodies, are also reported. In every case, structural changes associated with the point mutation are confined to the mutation site or to residues that are spatially immediately adjacent to it. The failure of antisera to cross-react between N2 and N9 subtypes may be correlated with the absence of conserved, contiguous surface structures of area 700 A2 or more. 相似文献
10.
Probing the Sialic Acid Binding Site of the Hemagglutinin-Neuraminidase of Newcastle Disease Virus: Identification of Key Amino Acids Involved in Cell Binding, Catalysis, and Fusion 总被引:5,自引:0,他引:5
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Helen Connaris Toru Takimoto Rupert Russell Susan Crennell Ibrahim Moustafa Allen Portner Garry Taylor 《Journal of virology》2002,76(4):1816-1824
We recently reported the first crystal structure of a paramyxovirus hemagglutinin-neuraminidase (HN) from Newcastle disease virus. This multifunctional protein is responsible for binding to cellular sialyl-glycoconjugate receptors, promotion of fusion through interaction with the second viral surface fusion (F) glycoprotein, and processing progeny virions by removal of sialic acid from newly synthesized viral coat proteins. Our structural studies suggest that HN possesses a single sialic acid recognition site that can be switched between being a binding site and a catalytic site. Here we examine the effect of mutation of several conserved amino acids around the binding site on the hemagglutination, neuraminidase, and fusion functions of HN. Most mutations around the binding site result in loss of neuraminidase activity, whereas the effect on receptor binding is more variable. Residues E401, R416, and Y526 appear to be key for receptor binding. The increase in fusion promotion seen in some mutants that lack receptor binding activity presents a conundrum. We propose that in these cases HN may be switched into a fusion-promoting state through a series of conformational changes that propagate from the sialic acid binding site through to the HN dimer interface. These results further support the single-site model and suggest certain residues to be important for the triggering of fusion. 相似文献
11.
Normal horse and guinea pig sera contain the glycoprotein inhibitor alpha 2-macroglobulin, which inhibits the infectivity and hemagglutinating activity of influenza A viruses of the H2 and H3 subtypes. In the current study, the presence of inhibitors of influenza A virus in pig and rabbit sera was investigated. Variants of influenza virus type A/Los Angeles/2/87(H3N2) that were resistant to horse, pig, or rabbit serum were isolated. Analysis of the variant viruses with anti-hemagglutinin (HA) monoclonal antibodies revealed that antigenic changes occurred with the development of serum inhibitor resistance. Characterization of the inhibitors in pig and rabbit sera by using periodate and receptor-destroying enzyme demonstrated that carbohydrate is an important constituent of the active portion of both inhibitor molecules and that sialic acid is involved in the interaction of the inhibitors with influenza virus HA. Nucleotide sequence analysis of the HA molecule revealed that the serum-resistant variants each acquired a different set of amino acid alterations. The multiply resistant variants maintained the original amino acid changes and acquired additional changes. Sequence modifications in the HA involved the conserved amino acids within the receptor binding site (RBS) at position 137 and the second-shell RBS residues at positions 155 and 186. Amino acid changes also occurred within antigenic site A (position 145) and directly behind the receptor binding pocket (position 220). Amino acid alterations resulted in the acquisition of a potential glycosylation site at position 128 and the loss of potential glycosylation sites at positions 246 and 248. The localization of the amino acid changes in HA1 to the region of the RBS supports the concept of serum inhibitors as receptor analogs. The unique set of mutations acquired by the serum inhibitor-resistant variants strongly suggests that horse, pig, and rabbit sera each contain distinct glycoprotein inhibitors of influenza A virus. 相似文献
12.
The crystal structure of the ternary complex of staphylococcal nuclease, Ca2+, and the inhibitor pdTp, refined at 1.65 A 总被引:20,自引:0,他引:20
The structure of a complex of staphylococcal nuclease with Ca2+ and deoxythymidine 3',5'-bisphosphate (pdTp) has been refined by stereochemically restrained least-squares minimization to a crystallographic R value of 0.161 at 1.65 A resolution. The estimated root-mean-square (rms) error in the coordinates is 0.16 A. The final model comprises 1082 protein atoms, one calcium ion, the pdTp molecule, and 82 solvent water molecules; it displays an rms deviation from ideality of 0.017 A for bond distances and 1.8 degrees for bond angles. The mean distance between corresponding alpha carbons in the refined and unrefined structures is 0.6 A; we observe small but significant differences between the refined and unrefined models in the turn between residues 27 and 30, the loop between residues 44 and 50, the first helix, and the extended strand between residues 112 and 117 which forms part of the active site binding pocket. The details of the calcium liganding and solvent structure in the active site are clearly shown in the final electron density map. The structure of the catalytic site is consistent with the mechanism that has been proposed for this enzyme. However, we note that two lysines from a symmetry-related molecule in the crystal lattice may play an important role in determining the geometry of inhibitor binding, and that only one of the two required calcium ions is observed in the crystal structure; thus, caution is advised in extrapolating from the structure of the complex of enzyme and inhibitor to that of enzyme and substrate. 相似文献
13.
Site-directed mutagenesis of a conserved hexapeptide in the paramyxovirus hemagglutinin-neuraminidase glycoprotein: effects on antigenic structure and function. 总被引:14,自引:7,他引:7
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The sequence NRKSCS constitutes the longest linear stretch in the amino acid sequence of the hemagglutinin-neuraminidase (HN) glycoprotein of the paramyxoviruses that is completely conserved among all viruses in the group. We have used site-directed mutagenesis and expression of the mutated HN protein of one member of the group, Newcastle disease virus, to explore the role of this highly conserved sequence in the structure and function of the protein. Any substitution introduced for each of four residues in the sequence, N-234, R-235, K-236, or S-237, results in a drastic decrease in neuraminidase activity relative to that of the wild-type protein. Only substitutions for the terminal serine residue in the sequence had comparatively little effect on this activity. These findings are consistent with prior computer-based predictions of protein secondary structure which had suggested that this domain corresponds to one in the beta-sheet propeller structure of the neuraminidase protein of influenza virus closest to the center of the sialic acid binding site and forms part of the enzyme active site. Four of the substitutions, N-234-->Y and K-236-->E, -->Q, and -->S, apparently cause a local alteration in the antigenic structure of the protein. This is evidenced by (i) the diminished recognition of the protein only by monoclonal antibodies thought to bind at the neuraminidase active site, among an extensive panel of conformation-specific antibodies, and (ii) the slower rate of migration in sodium dodecyl sulfate-polyacrylamide gel electrophoresis for all except the K-236-->Q mutation. One of the mutations, K-236-->S, completely abolishes the ability of the protein to promote cellular fusion when coexpressed with the fusion protein. The latter cannot be explained by a decrease in the relative hemadsorption activity of the protein and suggests that the globular head of the protein may contribute to this process beyond providing receptor recognition. 相似文献
14.
15.
甲型H1N1流感病毒是一种高度接触性急性呼吸道传染病,给人类健康造成了极大威胁。作为一个已经被证明的抗流感药物靶点,神经氨酸酶在流感病毒复制和传播中发挥重要作用,并且其活性中心的氨基酸组成高度保守。本研究中,我们使用分子对接技术筛选从中草药小分子数据库中筛选神经氨酸酶潜在的抑制剂,结果得到4个亲和力高于奥司他韦(Osel-tamivir)的化合物,并确定了他们的中草药来源。为从草药中提取,设计以及实验合成新的神经氨酸酶抑制剂提供了一定的依据和指导。 相似文献
16.
This is the first structural observation of a plant product showing high affinity for phospholipase A(2) and regulating the synthesis of arachidonic acid, an intermediate in the production of prostaglandins. The crystal structure of a complex formed between Vipera russelli phospholipase A(2) and a plant alkaloid aristolochic acid has been determined and refined to 1.7 A resolution. The structure contains two crystallographically independent molecules of phospholipase A(2) in the form of an asymmetric dimer with one molecule of aristolochic acid bound to one of them specifically. The most significant differences introduced by asymmetric molecular association in the structures of two molecules pertain to the conformations of their calcium binding loops, beta-wings, and the C-terminal regions. These differences are associated with a unique conformational behavior of Trp(31). Trp(31) is located at the entrance of the characteristic hydrophobic channel which works as a passage to the active site residues in the enzyme. In the case of molecule A, Trp(31) is found at the interface of two molecules and it forms a number of hydrophobic interactions with the residues of molecule B. Consequently, it is pulled outwardly, leaving the mouth of the hydrophobic channel wide open. On the other hand, Trp(31) in molecule B is exposed to the surface and moves inwardly due to the polar environment on the molecular surface, thus narrowing the opening of the hydrophobic channel. As a result, the aristolochic acid is bound to molecule A only while the binding site of molecule B is empty. It is noteworthy that the most critical interactions in the binding of aristolochic acid are provided by its OH group which forms two hydrogen bonds, one each with His(48) and Asp(49). 相似文献
17.
The binding properties of the H5N1 influenza virus neuraminidase as inferred from molecular modeling
The avian influenza H5N1 virus has emerged as an important pathogen, causing severe disease in humans and posing a pandemic
threat. Substrate specificity is crucial for the virus to obtain the ability to spread from avian to human. Therefore, an
investigation of the binding properties of ligands at the molecular level is important for understanding the catalytic mechanism
of the avian influenza virus neuraminidase and for designing novel and specific inhibitors of H5N1 neuraminidase. Based on
the available crystal structure of H5N1, we have characterized the binding properties between sialic acid, methyl 3’sialyllactoside,
methyl 6’sialyllactoside and the H5N1 influenza virus neuraminidase using molecular docking and molecular dynamics simulations.
Obtained molecular dynamics trajectories were analyzed in terms of ligand conformations, N1-ligand interactions, and in terms
of loop flexibility. It was found that in the N1-SA complex the sialic acid ring undergoes a transition from the B
2,5 to the 2
C
5 conformation. However, in the N1-3SL and N1-6SL complexes sialic acid remained in the distorted boat conformation. The obtained
results indicate that 3SL has only weak interactions with the 150-loop, whereas the N1-6SL complex shows strong interactions.
Most of the differences arise from the various conformations around the glycosidic linkage, between the sialic acid and galactose,
which facilitate the above interactions of 6SL with the enzyme, and as a consequence the interactions between the 150- and
430- loops. This finding suggests that the altered flexibility of loops in and around the active site is one of the reasons
why the avian N1 preferentially cleaves sialic acid from α-(2-3)-Gal glycoconjugates over α-(2-6)-Gal. These molecular modeling
results are consistent with available experimental results on the specificity of N1. 相似文献
18.
目的:从噬菌体呈现12肽库中筛选与流感病毒神经氨酸酶特异性结合的肽。方法:以甲三型流感病毒裂解疫苗原液为靶分子,经过3轮生物淘选,从噬菌体随机肽库中筛选与之结合的噬菌体。用ELISA方法鉴定噬菌体克隆与靶分子的结合力,用荧光方法测定噬菌体克隆对流感病毒A/Sydney/5/97(H3N2)神经氨酸酶的抑制活性。对筛选到的阳性克隆进行DNA序列测定并推导出相应的氨基酸序列。结果:经过3轮筛选后,42个噬菌体克隆与靶分子有高度亲和力,23个噬菌体克隆对流感病毒A/Sydney/5/97(H3N2)神经氨酸酶有抑制活性。对27个噬菌体克隆的测序结果表明,分别有10个和2个克隆的序列是一致的,其氨基酸序列分别为KSLSRHDHIHHH和WPRHHHSASVQT。结论:通过噬菌体肽库筛选到抑制流感病毒神经氨酸酶的12肽,为进一步研究对流感病毒神经氨酸酶有抑制活性的分子药物奠定了基础。 相似文献
19.
Structure of the lysosomal sphingolipid activator protein 1 by homology with influenza virus neuraminidase 总被引:1,自引:0,他引:1
M Potier 《Biochemical and biophysical research communications》1988,155(1):32-37
The sphingolipid activator protein 1 (SAP-1) increases the rate of hydrolysis of sphingolipids in the lysosome by apparently bringing together the substrate and the corresponding hydrolytic enzyme. This implies specific recognition of both the substrate and enzyme by SAP-1. However, binding domains in SAP-1 and recognition mechanisms involved are unknown. Amino acid sequence comparison of SAP-1 with influenza virus neuraminidase (EC 3.2.1.18, FLU NA) indicates that functional amino acid residues in or near the sialic acid binding site of FLU NA are also found at equivalent positions in the first 48 N-terminal amino acids of SAP-1. This region of homology allows to propose folding of the SAP-1 polypeptide chain by comparison with known crystallographic structure of FLU NA and identify a potential domain for lysosomal enzyme recognition through sialic acid binding. There is also a region of 10 amino acid residues near the C-terminal end of SAP-1 which has a strong propensity to form an alpha-helix with amphiphilic properties of lipid-binding helices. This domain in SAP-1 is probably responsible for the lipid(substrate)-binding function of SAP-1. 相似文献
20.
G D Glick P L Toogood D C Wiley J J Skehel J R Knowles 《The Journal of biological chemistry》1991,266(35):23660-23669
Infection by influenza virus is initiated by a cellular adhesion event that is mediated by the viral protein, hemagglutinin, which is exposed on the surface of the virion. Hemagglutinin recognizes and binds to cell surface sialic acid residues. Although each individual ligand binding interaction is weak, the high affinity of influenza virus for cells that bear sialic acid residues is thought to result from a multivalent attachment process involving many similar recognition events. To evaluate such binding we have synthesized three series of compounds, each containing two sialic acid residues separated by spacers of different length, and have tested them as ligands for influenza hemagglutinin. No increased binding to the bromelain-released hemagglutinin ectodomain was seen for any of the bivalent compounds as determined by 1H NMR titration. In contrast, however, a spacer length between sialic acid residues of approximately 55 A sharply increases the binding of these bidentate species to whole virus as determined by hemagglutination inhibition assays. The most effective compound containing glycines in the linking chain displayed 100-fold increased affinity for whole virus over the paradigm monovalent ligand, Neu5Ac alpha 2Me. 相似文献