Polypeptide binding of Escherichia coli FtsH (HflB)

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Abstract
The Escherichia coli FtsH protein is a membrane‐bound and ATP‐dependent protease. In this study, we describe ATP‐dependent conformational changes in FtsH as well as a polypeptide binding ability of this protein. A 33 kDa segment of FtsH became trypsin resistant in the presence of ATP. ATP and ATPγS prevented self‐aggregation of detergent‐solubilized FtsH‐His6‐Myc at 37°C, again suggesting that the binding of ATP induces a conformational change in FtsH. Affinity chromatography showed that FtsH‐His6‐Myc can associate with denatured alkaline phosphatase (PhoA) but not with the native enzyme. Denatured PhoA also prevented the aggregation of FtsH, and these two proteins co‐sedimented through a sucrose gradient. Binding between FtsH‐His6‐Myc and detergent‐solubilized SecY was also demonstrated. Although FtsH‐bound SecY was processed further for ATP‐dependent proteolysis, FtsH‐bound PhoA was not. Thus, FtsH association with denatured PhoA is uncoupled from proteolysis. Overproduction of FtsH significantly increased the cytoplasmic localization of the PhoA moiety of a MalF–PhoA hybrid protein, in which a charged residue had been introduced into a transmembrane segment. Thus, denatured PhoA binding of FtsH may also occur in vivo.