The nonstructural protein 8 (nsp8) of the SARS coronavirus interacts with its ORF6 accessory protein
Abstract
Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) caused a severe outbreak in several regions of the world in 2003. The SARS-CoV genome is predicted to contain 14 functional open reading frames (ORFs). The first ORF (1a and 1b) encodes a large polyprotein that is cleaved into nonstructural proteins (nsp). The other ORFs encode for four structural proteins (spike, membrane, nucleocapsid and envelope) as well as eight SARS-CoV-specific accessory proteins (3a, 3b, 6, 7a, 7b, 8a, 8b and 9b). In this report we have cloned the predicted nsp8 gene and the ORF6 gene of the SARS-CoV and studied their abilities to interact with each other. We expressed the two proteins as fusion proteins in the yeast two-hybrid system to demonstrate protein-protein interactions and tested the same using a yeast genetic cross. Further the strength of the interaction was measured by challenging growth of the positive interaction clones on increasing gradients of 2-amino trizole. The interaction was then verified by expressing both proteins separately in-vitro in a coupled-transcription translation system and by coimmunoprecipitation in mammalian cells. Finally, colocalization experiments were performed in SARS-CoV infected Vero E6 mammalian cells to confirm the nsp8-ORF6 interaction. To the best of our knowledge, this is themore »
- Authors:
-
- Virology Group, International Centre for Genetic Engineering and Biotechnology, P. O. Box: 10504, Aruna Asaf Ali Road, New Delhi 110067 (India)
- Collaborative Anti-Viral Research Group, Institute of Molecular and Cell Biology, 61 Biopolis Drive, Proteos, Singapore 138673 (Singapore)
- Microbiology Department, National University of Singapore, Kent Ridge, Singapore 117597 (Singapore)
- Victorian Infectious Diseases Reference Laboratory, North Melbourne, Victoria (Australia)
- Publication Date:
- OSTI Identifier:
- 21077969
- Resource Type:
- Journal Article
- Journal Name:
- Virology
- Additional Journal Information:
- Journal Volume: 366; Journal Issue: 2; Other Information: DOI: 10.1016/j.virol.2007.04.029; PII: S0042-6822(07)00314-5; Copyright (c) 2007 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0042-6822
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 60 APPLIED LIFE SCIENCES; ANIMAL GROWTH; GENES; IN VITRO; MEMBRANES; PROTEINS; TRANSCRIPTION; VIRUSES; YEASTS
Citation Formats
Kumar, Purnima, Gunalan, Vithiagaran, Boping, Liu, Chow, Vincent T.K., Druce, Julian, Birch, Chris, Catton, Mike, Fielding, Burtram C, Tan, Yee-Joo, and Lal, Sunil K. The nonstructural protein 8 (nsp8) of the SARS coronavirus interacts with its ORF6 accessory protein. United States: N. p., 2007.
Web. doi:10.1016/j.virol.2007.04.029.
Kumar, Purnima, Gunalan, Vithiagaran, Boping, Liu, Chow, Vincent T.K., Druce, Julian, Birch, Chris, Catton, Mike, Fielding, Burtram C, Tan, Yee-Joo, & Lal, Sunil K. The nonstructural protein 8 (nsp8) of the SARS coronavirus interacts with its ORF6 accessory protein. United States. https://doi.org/10.1016/j.virol.2007.04.029
Kumar, Purnima, Gunalan, Vithiagaran, Boping, Liu, Chow, Vincent T.K., Druce, Julian, Birch, Chris, Catton, Mike, Fielding, Burtram C, Tan, Yee-Joo, and Lal, Sunil K. 2007.
"The nonstructural protein 8 (nsp8) of the SARS coronavirus interacts with its ORF6 accessory protein". United States. https://doi.org/10.1016/j.virol.2007.04.029.
@article{osti_21077969,
title = {The nonstructural protein 8 (nsp8) of the SARS coronavirus interacts with its ORF6 accessory protein},
author = {Kumar, Purnima and Gunalan, Vithiagaran and Boping, Liu and Chow, Vincent T.K. and Druce, Julian and Birch, Chris and Catton, Mike and Fielding, Burtram C and Tan, Yee-Joo and Lal, Sunil K.},
abstractNote = {Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) caused a severe outbreak in several regions of the world in 2003. The SARS-CoV genome is predicted to contain 14 functional open reading frames (ORFs). The first ORF (1a and 1b) encodes a large polyprotein that is cleaved into nonstructural proteins (nsp). The other ORFs encode for four structural proteins (spike, membrane, nucleocapsid and envelope) as well as eight SARS-CoV-specific accessory proteins (3a, 3b, 6, 7a, 7b, 8a, 8b and 9b). In this report we have cloned the predicted nsp8 gene and the ORF6 gene of the SARS-CoV and studied their abilities to interact with each other. We expressed the two proteins as fusion proteins in the yeast two-hybrid system to demonstrate protein-protein interactions and tested the same using a yeast genetic cross. Further the strength of the interaction was measured by challenging growth of the positive interaction clones on increasing gradients of 2-amino trizole. The interaction was then verified by expressing both proteins separately in-vitro in a coupled-transcription translation system and by coimmunoprecipitation in mammalian cells. Finally, colocalization experiments were performed in SARS-CoV infected Vero E6 mammalian cells to confirm the nsp8-ORF6 interaction. To the best of our knowledge, this is the first report of the interaction between a SARS-CoV accessory protein and nsp8 and our findings suggest that ORF6 protein may play a role in virus replication.},
doi = {10.1016/j.virol.2007.04.029},
url = {https://www.osti.gov/biblio/21077969},
journal = {Virology},
issn = {0042-6822},
number = 2,
volume = 366,
place = {United States},
year = {Sun Sep 30 00:00:00 EDT 2007},
month = {Sun Sep 30 00:00:00 EDT 2007}
}