The Mechanosensitive channel of Small conductance (MscS) of E. coli. senses membrane tension and opens to prevent hypoosmotic shock to the bacterium . Cardiolipin is present in the inner membrane of E. coli and induction of cardiolipin biosynthesis has been linked to osmotic stress response in E. coli . Previous studies showed that this channel co-localizes with cardiolipin at the poles of the bacterium , and that cardiolipin interaction with MscS causes increase in closing frequency of the channel when the channel is reconstituted into proteoliposomes. However the physiological relevance of this phenomenon remains unclear. Furthermore, cholesterol has been shown to influence MscS properties . Preliminary molecular modelling studies have shown a strong interaction of residues R46 and L60 with cardiolipin. Electrophysiological results from mutagenesis studies in which these residues were replaced by alanine suggest that these positively charged side chains play an important role in channel gating by interacting with negatively charged cardiolipin which is reported in our study.