Expression and membrane integration of SARS-CoV M protein |
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Authors: | Hsin-Chieh Ma Chiu-Ping Fang Yi-Ching Hsieh Shih-Chi Chen Hui-Chun Li Shih-Yen Lo |
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Affiliation: | (1) Graduate Institute of Medical Sciences, Tzu Chi University, 701, Section 3, Chung Yang Road, Hualien, Taiwan;(2) Graduate Institute of Medical Biotechnology, Tzu Chi University, Hualien, Taiwan;(3) Graduate Institute of Molecular and Cellular Biology, Tzu Chi University, Hualien, Taiwan |
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Abstract: | SARS-CoV M gene fragment was cloned and expressed as a recombinant protein fused with a V5 tag at the C-terminus in Vero E6 cells. In addition to un-glycosylated and glycosylated proteins, one product with smaller size initiated in-frame from the third Met residues probably through ribosomal re-initiation was also detected. Translation initiated in-frame from the third Met is unusual since the sequence around the first Met of SARS-CoV M protein contains the optimal consensus Kozak sequence. The function of this smaller translated product awaits further investigation. Similar to other N-glycosylated proteins, glycosylation of SARS-CoV M protein was occurred co-translationally in the presence of microsomes. The SARS-CoV M protein is predicted as a triple-spanning membrane protein lack of a conventional signal peptide. The second and third trans-membrane regions (a.a. 46–68 and 78–100) are predicted to be the primary type helices, which will be able to penetrate into membrane by themselves, while the first trans-membrane region (a.a. 14–36) is predicted to be the secondary type helix, which is considered to be stabilized by the interaction with other trans-membrane segments. As expected, the second and third trans-membrane regions were able to insert a cytoplasmic protein into the endoplasmic reticulum membrane more efficiently than the first one. These results should be important for the study of SARS-CoV morphogenesis. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. |
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Keywords: | SARS-CoV Membrane protein E.R. association Primary type transmembrane helix Co-translational event |
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