Latest Publications

Cook Jonathan, Crow Allister

The FtsEX-EnvC-AmiA/B system is a key component of the E. coli cell division machinery that directs breakage of the peptidoglycan layer during separation of daughter cells. Structural and mechanistic studies have shown that ATP binding by FtsEX in the cytoplasm drives periplasmic conformational changes in EnvC, which lead to the binding and activation of peptidoglycan amidases such as AmiA and AmiB. The FtsEX-EnvC amidase system is highly regulated to prevent cell lysis with at least two separate layers of autoinhibition that must be relieved to initiate peptidoglycan hydrolysis during division. Here, we test the FtsEX-EnvC amidase activation mechanism through site-directed mutagenesis. We identify mutations that disrupt the autoinhibition mechanism of FtsEX-EnvC and an N-terminal deletion variant that prevents activation. Finally, we develop a cysteine locking residue pair that stabilizes the complex in its amidase activating conformation. The reported EnvC variants greatly enhance our understanding of the FtsEX-EnvC autoinhibition mechanism and the conformational changes underpinning amidase activation. Our observations are consistent with the proposed mechanism of amidase activation by large-scale conformational changes in FtsEX-EnvC, allowing recruitment and activation of peptidoglycan amidases.

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