Infection with yellow fever virus (YFV), the prototypic mosquito-borne flavivirus, causes severe febrile disease with haemorrhage, multi-organ failure and a high mortality. Moreover, in recent years ...the Flavivirus genus has gained further attention due to re-emergence and increasing incidence of West Nile, dengue and Japanese encephalitis viruses. Potent and safe antivirals are urgently needed.
Starting from the crystal structure of the NS3 helicase from Kunjin virus (an Australian variant of West Nile virus), we identified a novel, unexploited protein site that might be involved in the helicase catalytic cycle and could thus in principle be targeted for enzyme inhibition. In silico docking of a library of small molecules allowed us to identify a few selected compounds with high predicted affinity for the new site. Their activity against helicases from several flaviviruses was confirmed in in vitro helicase/enzymatic assays. The effect on the in vitro replication of flaviviruses was then evaluated.
Ivermectin, a broadly used anti-helminthic drug, proved to be a highly potent inhibitor of YFV replication (EC₅₀ values in the sub-nanomolar range). Moreover, ivermectin inhibited, although less efficiently, the replication of several other flaviviruses, i.e. dengue fever, Japanese encephalitis and tick-borne encephalitis viruses. Ivermectin exerts its effect at a timepoint that coincides with the onset of intracellular viral RNA synthesis, as expected for a molecule that specifically targets the viral helicase.
The well-tolerated drug ivermectin may hold great potential for treatment of YFV infections. Furthermore, structure-based optimization may result in analogues exerting potent activity against flaviviruses other than YFV.
Chikungunya virus, Cameroon, 2006 Peyrefitte, Christophe N; Rousset, Dominique; Pastorino, Boris A M ...
Emerging infectious diseases,
05/2007, Letnik:
13, Številka:
5
Journal Article
Recenzirano
Odprti dostop
We report the isolation of chikungunya virus from a patient during an outbreak of a denguelike syndrome in Cameroon in 2006. The virus was phylogenetically grouped in the Democratic Republic of the ...Congo cluster, indicating a continuous circulation of a genetically similar chikungunya virus population during 6 years in Central Africa.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes epidemic fever, rash and polyarthralgia in Africa and Asia. Although it is known since the 1950s, new epidemiological and clinical ...features reported during the recent outbreak in the Indian Ocean can be regarded as the emergence of a new disease. Numerous severe forms of the infection have been described that put emphasis on the lack of efficient antiviral therapy. Among the virus-encoded enzymes, nsP2 constitutes an attractive target for the development of antiviral drugs. It is a multifunctional protein of approximately 90kDa with a helicase motif in the N-terminal portion of the protein while the papain-like protease activity resides in the C-terminal portion. The nsP2 proteinase is an essential enzyme whose proteolytic activity is critical for virus replication.
In this work, a recombinant CHIKV nsP2pro and a C-terminally truncated variant were expressed in Escherichia coli and purified by metal–chelate chromatography. The enzymatic properties of the proteinase were then determined using specific synthetic fluorogenic substrates. This study constitutes the first characterization of a recombinant CHIKV nsP2 cysteine protease, which may be useful for future drug screening.
The genus
Flavivirus, family
Flaviviridae, comprises more than 70 viruses. Many of them cause severe, potentially fatal, human diseases. Human vaccines are available for only three viruses and no ...effective antiviral drug is available. In order to limit the consequences of infections with flaviviruses, a promising approach consists in developing specific compounds that target the virus-encoded NS2B/NS3 protease complex, which is crucial for the viral polyprotein processing. In order to develop such compounds active as antiviral drugs against several flaviviruses, identification of biochemical properties shared by proteases from different viruses is essential.
In this work, the functional similarity between the proteases from seven flaviviruses belonging to different major groups was addressed by characterizing their enzymatic properties. For each virus, a catalytically active recombinant protease was designed and expressed as a hexahistidine-tagged protein. Chromogenic and fluorogenic substrates were used to identify optimal conditions for proteolysis. Our study identified important physico-chemical properties shared by all the seven proteases we studied (high pH value requirement for optimal activity, inhibition of substrate processing by salt). However, it also evidenced slight differences in biochemical properties of the flaviviral proteases, which could sustain heterogeneous sensitivity to future inhibitors.
Dengue type 3 virus, Saint Martin, 2003-2004 Peyrefitte, Christophe N; Pastorino, Boris A M; Bessaud, Maël ...
Emerging infectious diseases,
05/2005, Letnik:
11, Številka:
5
Journal Article
Recenzirano
Odprti dostop
We describe the spread of a dengue virus during an outbreak in Saint Martin island (French West Indies) during winter 2003-2004. Dengue type 3 viruses were isolated from 6 patients exhibiting ...clinical symptoms. This serotype had not been detected on the island during the preceding 3 years. Genome sequence determinations and analyses showed a common origin with dengue type 3 viruses isolated in Martinique 2 years earlier.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
O'nyong-nyong Virus, Chad Bessaud, Maël; Peyrefitte, Christophe N; Pastorino, Boris A M ...
Emerging infectious diseases,
08/2006, Letnik:
12, Številka:
8
Journal Article
Recenzirano
Odprti dostop
We report the first laboratory-confirmed human infection with O'nyong-nyong virus in Chad. This virus was isolated from peripheral blood mononuclear cells of a patient with evidence of a ...seroconversion to a virus related to Chikungunya virus. Genome sequence was partly determined, and phylogenetic studies were conducted.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
We report herein the study of the cleavage fragments generated by autoproteolysis of the St. Louis encephalitis virus recombinant protease. The cleavage sites leading to truncated forms were ...identified by microsequencing, which revealed an unexpected altered specificity of the recombinant proteinase towards unusual sequences.
Cell fusing agent virus (CFAV) is a positive strand RNA insect virus first isolated from a mosquito cell line. Based on viral morphology, phenotypic and phylogenetic studies, CFAV had been ...tentatively assigned to the genus Flavivirus (family Flaviviridae). The determination of the CFAV polyprotein complete sequence showed a putative serine protease domain analogue to the flaviviral NS2B/NS3 complex. This complex had been extensively studied, because it represented one of the main targets for antiflavivirus therapy development. We report herein the biochemical characterization of CFAV ΔNS2B-NS3pro protease complex. CFAV polyprotein sequence was computationally analysed to identify the amino-acid regions involved in protease activity. We designed, expressed and purified a catalytically active protease whose enzymatic properties were determined using fluorogenic substrates. Our results showed that, despite the low level of conservation of its amino-acid sequence, CFAV protease exhibited physico-chemical properties of other flaviviruses (high pH value requirement for optimal activity, inhibition by salt and preference for substrates featuring a basic residue at P₁ position).
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