Binding interaction of SARS coronavirus 3CL(pro) protease with vacuolar-H+ ATPase G1 subunit.
The pathogenesis of severe acute respiratory syndrome coronavirus (SARS-CoV) is an important issue for treatment and prevention of SARS. Recently, SARS-CoV 3CL(pro) protease has been implied to be possible relevance to SARS-CoV pathogenesis. In this study, we intended to identify potential 3CL(pro)-interacting cellular protein(s) using the phage-displayed human lung cDNA library. The vacuolar-H+ ATPase (V-ATPase) G1 subunit that contained a 3CL(pro) cleavage site-like motif was identified as a 3CL(pro)-interacting protein, as confirmed using the co-immunoprecipitation assay and the relative affinity assay. In addition, our result also demonstrated the cleavage of the V-ATPase G1 fusion protein and the immunoprecipitation of cellular V-ATPase G1 by the 3CL(pro). Moreover, loading cells with SNARF-1 pH-sensitive dye showed that the intracellular pH in 3CL(pro)-expressing cells was significantly lower as compared to mock cells.
Lin
Cheng-Wen
CW
Department of Medical Laboratory Science and Biotechnology, China Medical University, No. 91, Hsueh-Shih Road, Taichung 404, Taiwan. cwlin@mail.cmu.edu.tw
Tsai
Fuu-Jen
FJ
Wan
Lei
L
Lai
Chien-Chen
CC
Lin
Kuan-Hsun
KH
Hsieh
Tsung-Han
TH
Shiu
Shi-Yi
SY
Li
Jeng-Yi
JY
D000595:Amino Acid Sequence; D001578:Benzopyrans / Q000378:metabolism; D000086782:Coronavirus 3C Proteases; D003546:Cysteine Endopeptidases; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D047468:Immunoprecipitation; D008168:Lung / Q000201:enzymology*; D008969:Molecular Sequence Data; D009284:Naphthols / Q000378:metabolism; D019151:Peptide Library; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012235:Rhodamines / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D001578:Benzopyrans; D009284:Naphthols; D019151:Peptide Library; D021122:Protein Subunits; D011994:Recombinant Proteins; D012235:Rhodamines; D014764:Viral Proteins; C091666:seminaphthorhodaminefluoride; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D000086782:Coronavirus 3C Proteases; D025262:Vacuolar Proton-Translocating ATPases