OBJECTIVETo analyze and predict the structure and function of mosquito densovirus (MDV) nostructual protein1 (NS1).

METHODSUsing different bioinformatics software, the EXPASY pmtparam tool, ClustalX1.83, Bioedit, MEGA3.1, ScanProsite, and Motifscan, respectively to comparatively analyze and predict the physic-chemical parameters, homology, evolutionary relation, secondary structure and main functional motifs of NS1.

RESULTSMDV NS1 protein was a unstable hydrophilic protein and the amino acid sequence was highly conserved which had a relatively closer evolutionary distance with infectious hypodermal and hematopoietic necrosis virus (IHHNV). MDV NS1 has a specific domain of superfamily 3 helicase of small DNA viruses. This domain contains the NTP-binding region with a metal ion-dependent ATPase activity. A virus replication roller rolling-circle replication(RCR) initiation domain was found near the N terminal of this protein. This protien has the biological function of single stranded incision enzyme.

CONCLUSIONThe bioinformatics prediction results suggest that MDV NS1 protein plays a key role in viral replication, packaging, and the other stages of viral life.

Animals ; Computational Biology ; Culicidae ; virology ; Densovirus ; chemistry ; classification ; genetics ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; Protein Structure, Secondary ; Protein Structure, Tertiary ; Viral Nonstructural Proteins ; chemistry ; genetics

The present study was aimed to isolate the active compounds from the fermentation products of Fusarium oxysporum, which had hepatitis C virus (HCV) NS3 protease inhibitory activity. A bioactive compound was isolated by reverse-phase silica-gel column chromatography, silica-gel column chromatography, semi-preparative reverse-phase High Performance Liquid Chromatography (HPLC), and then its molecular structure was elucidated based on the spectrosopic analysis. As a result, the compound (H1-A, 1) Ergosta-5, 8 (14), 22-trien-7-one, 3-hydroxy-,(3β, 22E) was isolated and identified. To the best of our knowledge, this was the first report on the isolation of H1-A from microorganisms with the inhibitory activity of NS3 protease.
Enzyme Inhibitors ; chemistry ; isolation & purification ; metabolism ; Fusarium ; chemistry ; metabolism ; Hepacivirus ; drug effects ; enzymology ; genetics ; Hepatitis C ; virology ; Humans ; Magnetic Resonance Spectroscopy ; Viral Nonstructural Proteins ; antagonists & inhibitors ; metabolism

The hepatitis C virus (HCV) RNA-dependent RNA polymerase, NS5B protein, is the key viral enzyme responsible for replication of the HCV viral RNA genome. Although several full-length and truncated forms of the HCV NS5B proteins have been expressed previously in insect cells, contamination of host terminal transferase (TNTase) has hampered analysis of the RNA synthesis initiation mechanism using natural HCV RNA templates. We have expressed the HCV NS5B protein in insect cells using a recombinant baculovirus and purified it to near homogeneity without contaminated TNTase. The highly purified recombinant HCV NS5B was capable of copying 9.6-kb full-length HCV RNA template, and mini-HCV RNA carrying both 5'- and 3'-untranslated regions (UTRs) of the HCV genome. In the absence of a primer, and other cellular and viral factors, the NS5B could elongate over HCV RNA templates, but the synthesized products were primarily in the double stranded form, indicating that no cyclic replication occurred with NS5B alone. RNA synthesis using RNA templates representing the 3'-end region of HCV minus-strand RNA and the X-RNA at the 3'-end of HCV RNA genome was also initiated de novo. No formation of dimersize self-primed RNA products resulting from extension of the 3'-end hydroxyl group was observed. Despite the internal de novo initiation from the X-RNA, the NS5B could not initiate RNA synthesis from the internal region of oligouridylic acid (U)20, suggesting that HCV RNA polymerase initiates RNA synthesis from the selected region in the 3'-UTR of HCV genome.
3' Untranslated Regions/genetics ; 5' Untranslated Regions/genetics ; Animals ; Cell Line ; Gene Expression ; Genome ; Genome, Viral ; Hepacivirus/*enzymology/genetics ; RNA/biosynthesis/genetics ; RNA, Viral/genetics/metabolism ; Recombinant Proteins/genetics/isolation & purification/metabolism ; Spodoptera ; Templates, Genetic ; Uridine Monophosphate/metabolism ; Viral Nonstructural Proteins/chemistry/*genetics/isolation & purification/*metabolism

In order to investigate the transmission risk of H5N1 avian influenza viruses (AIV) from sewage in Changsha poultry markets, the evolution relationship and molecular characteristics of non-structural (NS) genes of H5N1 AIV from sewage were analyzed. Nine H5N1 AIV environmental sewage specimens were collected from Changsha poultry markets. The NS genes were amplifyed by PCR and then sequenced with TA cloning. Amino acid(aa) sequence alignment and phylogenetic tree analysis were conducted by Lasergene and Mega5 software. Eight NS genes TA cloning were constructed successfully. Phylogenetic tree indicated that they were belonged to the allele A subgroup. Aa homology analysis showed 90.1% 92.5% identity in NS1 proteins and 91.0% - 92.6% identity in NS2 proteins compared with reference viruses of the allele A (A/chicken/ Hubei/ w h/ 1999). The homologies of the amino sequences of NS1 and NS2 in this study were 93.8%-100.0% and 98.4%-100.0%, respectively. The C terminal of all eight H5N1 NS1 proteins from sewage in poultry markets carried a ESEV of PL motif and the 92 amino acids were E, furthermore, the 80 to 84aa were missed which were the characteristics of highly pathogenic AIV. The NS genes of H5N1 AIV from sewage in poultry markets have molecular characteristics of highly pathogenic and have the potential risk of H5N1 virus spreading.
Amino Acid Sequence ; Animals ; Influenza A Virus, H5N1 Subtype ; chemistry ; classification ; genetics ; isolation & purification ; Influenza in Birds ; transmission ; virology ; Molecular Sequence Data ; Phylogeny ; Poultry ; Sequence Homology, Amino Acid ; Sewage ; virology ; Viral Nonstructural Proteins ; chemistry ; genetics

To develop a sensitive and specific ELISA for detection of antibodies to the nonstructural protein of FMDV. We cloned and expressed FMDV nonstructural protein 3AB in Escherichia coli expression system. The recombinant protein 3AB was purified with Ni-NTA HisBind Resins and characterized by Western blotting. An indirect ELISA based on purified protein 3AB as a coating antigen was established. The specificity and sensitivity of this assay were evaluated by comparison with a commercial 3ABC-ELISA kit in detecion of serum samples. The results showed that the recombinant protein 3AB was expressed as a formation of inclusion bodies in Escherichia coli. The purified protein could specificially react with FMDV infection antibodies in Western blotting assay, but no reaction with the immune antibodies induced with vaccine. Two assays were no significant differences in specificity and sensitivity for detection of field samples (P>0.05). Therefore, we speculated that the recombinant protein 3AB is a promising molecular marker, which may effectively differentiate FMD-infected from vaccinated animals in a herd.
Animals ; Antibodies, Viral ; analysis ; Antigens, Viral ; biosynthesis ; genetics ; immunology ; Cattle ; Cattle Diseases ; diagnosis ; immunology ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; genetics ; metabolism ; Foot-and-Mouth Disease ; diagnosis ; immunology ; Foot-and-Mouth Disease Virus ; chemistry ; genetics ; isolation & purification ; Genetic Vectors ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Viral Nonstructural Proteins ; biosynthesis ; genetics ; immunology

Human enterovirus 71 (EV71) is one of the major etiological agents for the hand, foot, and month disease (HFMD) and is causing frequent, widespread occurrence in the mainland of China. The single positive-stranded RNA genome of EV71 is translated into a single polyprotein which is autocleavaged into structural and nonstructural proteins. The functions of many nonstructural proteins characterized in the life cycle of virus are potential targets for blocking viral replication. This article reviews the studies of the structures and functions of nonstructural proteins of EV71 and the anti-enterovirus 71 drugs targeting on these nonstructural proteins.
Antiviral Agents ; pharmacology ; Enterovirus A, Human ; enzymology ; genetics ; isolation & purification ; Hand, Foot and Mouth Disease ; drug therapy ; virology ; Humans ; Molecular Targeted Therapy ; Peptide Hydrolases ; chemistry ; metabolism ; physiology ; Protein Kinase Inhibitors ; pharmacology ; RNA, Viral ; genetics ; Viral Nonstructural Proteins ; chemistry ; metabolism ; physiology ; Virus Replication ; drug effects