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Journal of Virology, September 2005, p. 12081-12087, Vol. 79, No. 18
0022-538X/05/$08.00+0     doi:10.1128/JVI.79.18.12081-12087.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

The 41-Amino-Acid C-Terminal Region of the Hepatitis E Virus ORF3 Protein Interacts with Bikunin, a Kunitz-Type Serine Protease Inhibitor

Shweta Tyagi,{dagger} Milan Surjit, and Sunil K. Lal*

Virology Group, International Centre for Genetic Engineering & Biotechnology, Aruna Asaf Ali Road, New Delhi 110067, India

Received 22 January 2005/ Accepted 5 June 2005

Hepatitis E virus (HEV), a human plus-stranded RNA virus, contains three open reading frames (ORF). Of these, ORF1 encodes the viral nonstructural polyprotein, ORF2 encodes the major capsid protein, and ORF3 codes for a phosphoprotein of undefined function. Recently, using the yeast two-hybrid system to screen a human cDNA liver library, we have isolated and characterized AMBP ({alpha}1-microglobulin/bikunin precursor), which specifically interacts with the ORF3 protein of HEV. The ORF3 protein expedites the processing and secretion of {alpha}1-microglobulin. When checked individually for interaction, the second processed protein from AMBP, bikunin, strongly interacted with the full-length ORF3 protein. This protein-protein interaction has been validated by immunoprecipitation in both COS-1 and Huh7 cells and by His6 pull-down assays. In dual-labeling immunofluorescent staining, followed by fluorescence microscopy of transfected human liver cells, ORF3 colocalized with endogenously expressed bikunin. Finally, a 41-amino-acid C-terminal region of ORF3 has been found to be responsible for interacting with bikunin. The importance of this virus-host protein-protein interaction, with reference to the viral life cycle, has been discussed.


* Corresponding author. Mailing address: Virology Group, International Centre for Genetic Engineering & Biotechnology, P. O. Box 10504, Aruna Asaf Ali Road, New Delhi 10067, India. Phone: 91 (11) 26176680. Fax: 91 (11) 26162316. E-mail: sunillal{at}icgeb.res.in.

{dagger} Present address: Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724


Journal of Virology, September 2005, p. 12081-12087, Vol. 79, No. 18
0022-538X/05/$08.00+0     doi:10.1128/JVI.79.18.12081-12087.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.




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