Expression of nonstructural protein NS3 of African horsesickness virus (AHSV): evidence for a cytotoxic effect of NS3 in insect cells, and characterization of the gene products in AHSV infected Vero cells

1995 ◽  
Vol 140 (2) ◽  
pp. 289-306 ◽  
Author(s):  
V. van Staden ◽  
M. A. Stoltz ◽  
H. Huismans
Keyword(s):  
Cytotoxic Effect ◽  
Insect Cells ◽  
Nonstructural Protein ◽  
Vero Cells ◽  
Gene Products ◽  
Nonstructural Protein Ns3 ◽  
African Horsesickness Virus

scholarly journals Variation of African horsesickness virus nonstructural protein NS3 in southern Africa

2001 ◽  
Vol 82 (1) ◽  
pp. 149-158 ◽  
Author(s):  
M. van Niekerk ◽  
V. van Staden ◽  
A. A. van Dijk ◽  
H. Huismans
Keyword(s):  
Southern Africa ◽  
Nonstructural Protein ◽  
Live Attenuated Vaccine ◽  
Field Isolates ◽  
Ns3 Protein ◽  
Current Vaccine ◽  
Outer Capsid ◽  
Reference Strains ◽  
Nonstructural Protein Ns3 ◽  
African Horsesickness Virus

NS3 protein sequences of recent African horsesickness virus (AHSV) field isolates, reference strains and current vaccine strains in southern Africa were determined and compared. The variation of AHSV NS3 was found to be as much as 36·3% across serotypes and 27·6% within serotypes. NS3 proteins of vaccine and field isolates of a specific serotype were found to differ between 2·3% and 9·7%. NS3 of field isolates within a serotype differed up to 11·1%. Our data indicate that AHSV NS3 is the second most variable AHSV protein, the most variable being the major outer capsid protein, VP2. The inferred phylogeny of AHSV NS3 corresponded well with the described NS3 phylogenetic clusters. The only exception was AHSV-8 NS3, which clustered into different groups than previously described. No obvious sequence markers could be correlated with virulence. Our results suggest that NS3 sequence variation data could be used to distinguish between field isolates and live attenuated vaccine strains of the same serotype.

scholarly journals Purification and Characterization of West Nile Virus Nucleoside Triphosphatase (NTPase)/Helicase: Evidence for Dissociation of the NTPase and Helicase Activities of the Enzyme

2001 ◽  
Vol 75 (7) ◽  
pp. 3220-3229 ◽  
Author(s):  
Peter Borowski ◽  
Andreas Niebuhr ◽  
Oliver Mueller ◽  
Maria Bretner ◽  
Krzysztof Felczak ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Atp Hydrolysis ◽  
Nonstructural Protein ◽  
Kinetic Studies ◽  
Immunoblot Analysis ◽  
Vero Cells ◽  
Helicase Activity ◽  
West Nile ◽  
Nucleoside Triphosphatase

ABSTRACT The nucleoside triphosphatase (NTPase)/helicase associated with nonstructural protein 3 of West Nile (WN) virus was purified from cell culture medium harvested from virus-infected Vero cells. The purification procedure included sequential chromatography on Superdex-200 and Reactive Red 120 columns, followed by a concentration step on an Ultrogel hydroxyapatite column. The nature of the purified protein was confirmed by immunoblot analysis using a WN virus-positive antiserum, determination of its NH2 terminus by microsequencing, and a binding assay with 5′-[14C]fluorosulfonylbenzoyladenosine. Under optimized reaction conditions the enzyme catalyzed the hydrolysis of ATP and the unwinding of the DNA duplex with k cat values of 133 and 5.5 × 10−3 s−1, respectively. Characterization of the NTPase activity of the WN virus enzyme revealed that optimum conditions with respect to the Mg2+requirement and the monovalent salt or polynucleotide response differed from those of other flavivirus NTPases. Initial kinetic studies demonstrated that the inhibition (or activation) of ATPase activity by ribavirin-5′-triphosphate is not directly related to changes in the helicase activity of the enzyme. Further analysis using guanine andO 6-benzoylguanine derivatives revealed that the ATPase activity of WN virus NTPase/helicase may be modulated, i.e., increased or reduced, with no effect on the helicase activity of the enzyme. On the other hand the helicase activity could be modulated without changing the ATPase activity. Our observations show that the number of ATP hydrolysis events per unwinding cycle is not a constant value.

scholarly journals Membrane Association of African Horsesickness Virus Nonstructural Protein NS3 Determines Its Cytotoxicity

Virology ◽  
2001 ◽  
Vol 279 (2) ◽  
pp. 499-508 ◽  
Author(s):  
Michelle van Niekerk ◽  
Carel C. Smit ◽  
Wilma C. Fick ◽  
Vida van Staden ◽  
Henk Huismans
Keyword(s):  
Nonstructural Protein ◽  
Membrane Association ◽  
Nonstructural Protein Ns3 ◽  
African Horsesickness Virus

Phytochemical screening, cytotoxicity and anti-inflammatory activities of the leaf extracts from Lawsonia inermis of Indian origin to explore its potential for medicinal uses

2020 ◽  
Vol 16 ◽  
Author(s):  
Anju Manuja ◽  
Nitu Rathore ◽  
Shalki Chaudhary ◽  
Balvinder Kumar
Keyword(s):  
Nitric Oxide ◽  
Cytotoxic Activity ◽  
Cytotoxic Effect ◽  
Vero Cells ◽  
Nitric Oxide Production ◽  
Lawsonia Inermis ◽  
Leaf Extracts ◽  
Oxide Production ◽  
Indian Origin ◽  
Anti Inflammatory

Background: Lawsonia inermis Linn popularly known as the Henna has played an important role in ayurvedic or natural herbal medicines. The presence of phyto-constituents in henna, that may affect the animal or human health adversely, need to be elucidated for L. inermis Linn species grown in India. Introduction: Introduction: The aim of this research was to perform phytochemical, cytotoxicity and anti-inflammatory studies to understand the potential of leaves of Lawsonia inermis of Indian origin to provide a way forward for therapeutic use in medicine. Methods: We assessed the phytochemical profile for presence of phyto-constituents (alkaloids, carbohydrates, glycosides, steroids, flavonoids, saponins, tannins, proteins/amino acids and gums/mucilage) from various extracts of the plant leaves’. The extracts were further purified by column chromatography for the isolation of plant constituents and monitored by TLC, analyzed by Fourier transform infrared FT-IR spectroscopy, H1NMR, and GC-MS analysis. Fractions were assessed for cytotoxicity and anti-inflammatory properties at various concentrations. We assessed the anti-inflammatory activity by nitric oxide production in various leaf extracts determined by Griess assay. Results: All the spectral results suggest that the compounds from the extract contain aromatic nucleus and OH group along with methoxy group, allyl as well as vinyl group. Fractions of chloroform/methanolic (7:3) leaf extract of Lawsonia inermis confirmed the presence of the two constituents i.e. fraxetin and 1(3H)-isobenzofuranone. We observed significant difference in cytotoxicity at higher concentrations in methanol and chloroform:methanol (8:2) leaf extracts (p>0.05), we could not find any significant differences amongst other leaf extracts at different concentrations. Some leaf extracts have potential cytotoxic activity on vero cells. Reducing the chloroform concentration during extraction decreases the cytotoxic effect on the cells. The nitric oxide levels decreased from 1000 µg/ml concentration to lower concentrations with varying degree. Overall the highest nitric oxide production by CHCl3 (70%)/ MeOH (30%) was observed amongst various fractions at different concentrations. Conclusion: The phytochemical, cytotoxicity and anti-inflammatory studies indicating the potential of leaves of the plant to provide a way further for their use in medicine. Fraxetin 1(3H)-isobenzofuranone structures were confirmed in fractions of CHCl3 (70%)/ MeOH (30%) extract as observed as a potent constituents. Some leaf extracts have potential cytotoxic activity on vero cells. Reducing the chloroform concentration during extraction decreases the cytotoxic effect on the cells.The cytotoxicity studies indicates the presence of cytotoxic compounds in some of these extracts, warranting research for fabrication of suitable formulations comprising these constituents to reduce its dose/toxicity for the use of beneficial effects of the plant components.

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