Development of a yeast two-hybrid screen for selection of A/H1N1 influenza NS1 non-structural protein and human CPSF30 protein interaction inhibitors / 药学学报
Yao Xue Xue Bao
; (12): 388-394, 2010.
Article
de Zh
| WPRIM
| ID: wpr-250574
Bibliothèque responsable:
WPRO
ABSTRACT
Influenza A/H1N1 virus-encoded nonstructural, or NS1, protein inhibits the 3'-end processing of cellular pre-mRNAs by binding the cellular protein: the 30-kDa subunit of CPSF (cleavage and polyadenylation specificity factor, CPSF30). CPSF30 binding site of the NS1 protein is a potential target for the development of drugs against influenza A/H1N1 virus. A yeast two-hybrid screening system was constructed and used for screening Chinese medicines that inhibit the interaction of the A/H1N1 flu NS1 protein and human CPSF30 protein. The NS1 gene of A/H1N1 virus was amplified by consecutive polymerase chain reaction (PCR), and the human CPSF30 gene of HeLa cell cloned by reverse transcriptase-polymerase chain reaction (RT-PCR). Then the two gene fragments confirmed by sequencing were subcloned into the yeast expression vectors pGBKT7 and pGADT7, respectively. The two constructs, bait vector pGBKNS1 and prey vector pGADCPSF, were co-transformed into yeast AH109. The eight individual yeast colonies were picked and subjected to verification by PCR/gel electrophoresis. The inhibition of the NS1-CPSF30 interaction was allowed the identification of selective inhibitors. The four of more than thirty identified Chinese medicines, including 'Shuanghuanglian oral liquid', showed the strong inhibition of the NS1-CPSF30 interaction.
Texte intégral:
1
Indice:
WPRIM
Sujet Principal:
Fragments peptidiques
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Pharmacologie
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Plasmides
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Liaison aux protéines
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Transformation génétique
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Sites de fixation
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Médicaments issus de plantes chinoises
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Séquence nucléotidique
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Cellules HeLa
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Amplification de gène
Type d'étude:
Prognostic_studies
Limites du sujet:
Humans
langue:
Zh
Texte intégral:
Yao Xue Xue Bao
Année:
2010
Type:
Article