Chak Sangma

An Avian Influenza H5N1 Virus That Binds to a Human-Type Receptor

ABSTRACT Avian influenza viruses preferentially recognize sialosugar chains terminating in sialic acid-α2,3-galactose (SAα2,3Gal), whereas human influenza viruses preferentially recognize SAα2,6Gal. A conversion to SAα2,6Gal specificity is believed to be one of the changes required for the introduction of new hemagglutinin (HA) subtypes to the human population, which can lead to pandemics. Avian influenza H5N1 virus is a major threat for the emergence of a pandemic virus. As of 12 June 2007, the virus has been reported in 45 countries, and 312 human cases with 190 deaths have been confirmed. We describe here substitutions at position 129 and 134 identified in a virus isolated from a fatal human case that could change the receptor-binding preference of HA of H5N1 virus from SAα2,3Gal to both SAα2,3Gal and SAα2,6Gal. Molecular modeling demonstrated that the mutation may stabilize SAα2,6Gal in its optimal cis conformation in the binding pocket. The mutation was found in approximately half of the viral sequences directly amplified from a respiratory specimen of the patient. Our data confirm the presence of H5N1 virus with the ability to bind to a human-type receptor in this patient and suggest the selection and expansion of the mutant with human-type receptor specificity in the human host environment.

Potent antitumor activity of synthetic 1,2-Naphthoquinones and 1,4-Naphthoquinones

Rhinacanthone (1) and two 1,2-pyranonaphthoquinones (2,3) were synthesized and found to show very potent cytotoxicity against three cancer cell lines (KB, HeLa and HepG(2)) with IC(50) values of 0.92-9.63 microM, whereas the corresponding hydroxylated derivative 4 had reduced cytotoxicity (IC(50) values of 7.61-24.13 microM). Three 1,2-furanonaphthoquinone derivatives (5-7) were also synthesized with similar cytotoxicity as 1,2-pyranonaphthoquinones. In comparison to 1,2-naphthoquinones, six 1,4-naphthoquinones derivatives fused with pyran ring (8-10) and furan ring (11-13) were synthesized and they showed less cytotoxicity or inactive to the cancer cell lines. Moreover, compound 13 had significant cytotoxicity against HeLa cell line (IC(50) value of 9.25 microM) while it showed no toxic to vero cell.

Influenza A virus molecularly imprinted polymers and their application in virus sub-type classification

In this work, we apply a molecular imprinting strategy as a screening protocol for different influenza A subtypes, namely H5N1, H5N3, H1N1, H1N3 and H6N1. Molecularly imprinted polymers for each of these subtypes lead to appreciable sensor characteristics on a quartz crystal microbalance leading to detection limits as low as 105 particles per ml. Selectivity studies indicate that each virus is preferably incorporated by its own MIP. Recognition in most cases is dominated by the neuraminidase residue rather than the hemagglutinin. Multivariate analysis shows that the sensor responses can be correlated with the differences in hemagglutinin and neuraminidase patterns from databases. This allows for virus subtype characterization and thus rapid screening.

Virtual screening for anti-HIV-1 RT and anti-HIV-1 PR inhibitors from the Thai medicinal plants database: a combined docking with neural networks approach.

The virtual screening approach for docking small molecules into a known protein structure is a powerful tool for drug design. In this work, a combined docking and neural network approach, using a self-organizing map, has been developed and applied to screen anti-HIV-1 inhibitors for two targets, HIV-1 RT and HIV-1 PR, from active compounds available in the Thai Medicinal Plants Database. Based on nevirapine and calanolide A as reference structures in the HIV-1 RT binding site and XK-263 in the HIV-1 PR binding site, 2,684 compounds in the database were docked into the target enzymes. Self-organizing maps were then generated with respect to three types of pharmacophoric groups. The map of the reference structures were then superimposed on the feature maps of all screened compounds. Only the structures having similar features to the reference compounds were accepted. By using the SOMs, the number of candidates for HIV-1 RT was reduced to six and nine compounds consistent with nevirapine and calanolide A, respectively, as references. For the HIV-1 PR target, there are 135 screened compounds showed good agreement with the XK-263 feature map. These screened compounds will be further tested for their HIV-1 inhibitory affinities. The obtained results indicate that this combined method is clearly helpful to perform the successive screening and to reduce the analyzing step from AutoDock and scoring procedure.

Prediction of avian influenza A binding preference to human receptor using conformational analysis of receptor bound to hemagglutinin

BackgroundIt is known that the highly pathogenic avian influenza A virus H5N1 binds strongly and with high specificity to the avian-type receptor by its hemagglutinin surface protein. This specificity is normally a barrier to viral transmission from birds to humans. However, strains may emerge with mutated hemagglutinin, potentially changing the receptor binding preference from avian to human-type. This hypothesis has been proven correct, since viral isolates from Vietnam and Thailand have been found which have increased selectivity toward the human cell receptor. The change in binding preference is due to mutation, which can be computationally modelled. The aim of this study is to further explore whether computational simulation could be used as a prediction tool for host type selectivity in emerging variants.ResultsMolecular dynamics simulation was employed to study the interactions between receptor models and hemagglutinin proteins from H5N1 strains A/Duck/Singapore/3/97, mutated A/Duck/Singapore/3/97 (Q222L, G224S, Q222L/G224S), A/Thailand/1(KAN-1)/2004, and mutated A/Thailand/1(KAN-1)/2004 (L129V/A134V). The avian receptor was represented by Siaα(2,3)Gal substructure and human receptor by Siaα(2,6)Gal. The glycoside binding conformation was monitored throughout the simulations since high selectivity toward a particular host occurs when the sialoside bound with the near-optimized conformation.ConclusionThe simulation results showed all hemagglutinin proteins used the same set of amino acid residues to bind with the glycoside; however, some mutations alter linkage preferences. Preference toward human-type receptors is associated with a positive torsion angle, while avian-type receptor preference is associated with a negative torsion angle. According to the conformation analysis of the bound receptors, we could predict the relative selectivity in accordance with in vitro experimental data when disaccharides receptor analogs were used.

A novel approach to identify molecular binding to the influenza virus H5N1: screening using molecularly imprinted polymers (MIPs)

In this report we investigate whether a molecularly imprinted polymer (MIP) of an inactivated strain of influenza A H5N1 could be used to help identify molecules capable of binding to, and inhibiting the function of the virus, via either competitive or allosteric mechanisms. Molecules which bind to the virus and induce a conformational change are expected to show reduced binding to the H5N1 specific MIP. Given the importance of molecular recognition in virus replication, such conformational change might also reduce the effectiveness of neuraminidase (N1) for cleaving the sialic groups necessary for virus replication. We show that the method can indeed differentiate between a potent neuraminidase inhibitor, H1 and H5 antibodies, and N1 specific and non-specific monosaccharide substrates. We suggest that such a method could potentially be used in conjunction with traditional biochemical assays to facilitate the identification of molecules functioning via novel modes of action.

A novel method for dengue virus detection and antibody screening using a graphene-polymer based electrochemical biosensor

Dengue fever is a major disease that kills many people in the developing world every year. During early infection, a patient displays a high temperature without other signs. After this stage, and without proper treatment, serious damage to internal organs can happen, which occasionally leads to death. A rapid technique for the early detection of dengue virus (DENV) could reduce the number of fatalities. This study presents a new technique for the detection, classification and antibody screening of DENV based on electrochemical impedance spectroscopy (EIS). We found that the charge transfer resistance (Rct) of a gold electrode coated with graphene oxide reinforced polymer was influenced by virus type and quantity exposed on the surface. Molecular recognition capability established during the GO-polymer composite preparation was used to explain this observation. The linear dependence of Rct versus virus concentrations ranged from 1 to 2×103pfu/mL DENV with a 0.12 pfu/mL detection limit.

In silico screening of epidermal growth factor receptor (EGFR) in the tyrosine kinase domain through a medicinal plant compound database

The unregulated epidermal growth factor receptor tyrosine kinase (ErbB1-TK or EGFR-TK) protein is involved in the proliferation of more than 50% of all cancer types. The reduction of EGFR-TK activity by small or medium-sized molecules has been proven to be an effective treatment for cancer. There is a widespread belief that Chinese medicinal herbs are active against several diseases, including various types of cancer. In this study, 29,960 compounds from the Chemiebase medicinal compound database were virtually screened against the EGFR-TK using AutoDock4.0, GOLD and GLIDE (XP). The results revealed eight potential hits: CAS nos. 104096-45-9, 112649-21-5, 113866-89-0, 142608-98-8, 142608-99-9, 144761-33-1, 155233-17-3 and 80510-05-0. These compounds have been reported to show anticancer activities in the literature. With the help of SiMMap and MOE interaction analysis, the protein–ligand interaction patterns between the functional groups of these compounds and the binding pocket residues were analyzed. Hydrogen bonding and hydrophobic forces are the main components of the interactions of these hits, similar to those observed for the known inhibitors erlotinib, gefitinib and AEE. The physicochemical filter indicates that compounds CAS nos. 104096-45-9 and 144761-33-1 are likely to be potential leads in the drug discovery process.

Receptor recognition mechanism of human influenza A H1N1 (1918), avian influenza A H5N1 (2004), and pandemic H1N1 (2009) neuraminidase.

Influenza A neuraminidase (NA) is a target for anti-influenza drugs. The function of this enzyme is to cleave a glycosidic linkage of a host cell receptor that links sialic acid (Sia) to galactose (Gal), to allow the virus to leave an infected cell and propagate. The receptor is an oligosaccharide on the host cell surface. There are two types of oligosaccharide receptor; the first, which is found mainly on avian epithelial cell surfaces, links Sia with Gal by an α2,3 glycosidic linkage; in the second, found mainly on human epithelial cell surfaces, linkage is via an α2,6 linkage. Some researchers believe that NAs from different viruses show selectivity for each type of linkage, but there is limited information available to confirm this hypothesis. To see if the linkage type is more specific to any particular NA, a number of NA-receptor complexes of human influenza A H1N1 (1918), avian influenza A H5N1 (2004), and a pandemic strain of H1N1 (2009) were constructed using homology modeling and molecular dynamics simulation. The results show that the two types of receptor analogues bound to NAs use different mechanisms. Moreover, it was found that a residue unique to avian virus NA is responsible for the recognition of the Siaα2,3Gal receptor, and a residue unique to human virus NA is responsible for the recognition of Siaα2,6Gal. We believe that this finding could explain how NAs of different virus origins always possess some unique residues.

Intra-host diversities of the receptor-binding domain of stork faeces-derived avian H5N1 viruses and its significance as predicted by molecular dynamic simulation.

Virus evolution facilitates the emergence of viruses with unpredictable impacts on human health. This study investigated intra-host variations of the receptor-binding domain (RBD) of the haemagglutinin (HA) gene of the avian H5N1 viruses obtained from the 2004 and 2005 epidemics. The results showed that the mutation frequency of the RBD ranged from 0.3 to 0.6 %. The mutations generated one consensus and several minor populations. The consensus population of the 2004 epidemic was transmitted to the 2005 outbreak with increased frequency (39 and 45 %, respectively). Molecular dynamics simulation was applied to predict the significance of the variants. The results revealed that the consensus sequence (E218K/V248I) interacted unstably with sialic acid (SA) with an α2,6 linkage (SAα2,6Gal). Although the mutated K140R/E218K/V248I and Y191C/E218K/V248I sequences decreased the HA binding capacity to α2,3-linked SA, they were shown to bind α2,6-linked SA with increased affinity. Moreover, the substitutions at aa 140 and 191 were positive-selection sites. These data suggest that the K140R and Y191C mutations may represent a step towards human adaptation of the avian H5N1 virus.