Regulation of potassium pool is essential for survival of bacteria submitted to hyperosmotic conditions. The potassium efflux systems KefA and KefD are believed to play a major role in regulating the potassium pool in Escherichia coli. The KefA system has been identified and cloned in the laboratory of Professor Booth in Aberdeen. The kefA locus has been sequenced and encodes three possible open reading frames. We will analyse the function of the three ORFs to determine which are responsible for KefA activity. The kefA products will be characterized in term of topology, cellular localization and activity. The regulation of ketA transcription and tlanslation will be studied by using gene fusion technology and primer extension. The influence of several factors on the activity of kefA, such as genetic background, medium osmolarity, growth phase and osmoprotectants presence will be investigated. The physiologic role of kefA will be studied by using defined kefA mutations created by reverse genetics.