Binding interaction of SARS coronavirus 3CLpro 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 3CLpro protease has been implied to be possible relevance to SARS-CoV pathogenesis. In this study, we intended to identify potential 3CLpro-interacting cellular protein(s) using the phage-displayed human lung cDNA library. The vacuolar-H+ ATPase (V-ATPase) G1 subunit that contained a 3CLpro cleavage site-like motif was identified as a 3CLpro-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 3CLpro. Moreover, loading cells with SNARF-1 pH-sensitive dye showed that the intracellular pH in 3CLpro-expressing cells was significantly lower as compared to mock cells.