The synthesis and hydrolysis of adenosine triphosphate (ATP), a biological energy carrier, as performed by F-type ATP synthases is an integral part to life. Structurally, much is known about the enzyme; especially with the recent advancement of high resolution cryo electron microscopy structures. Despite this information, the intricacies of the rotation mechanism occurring between subunits a and c are not fully understood. This rotation is ultimately responsible for utilizing the proton gradient across a membrane to synthesize ATP. Previous research identified specific residues of the c subunit of E. coli F-ATPase that may be necessary for function. The arginine at position 50 appears to be involved in key interactions due to either its steric bulk or its positive charge. This residue is essential to the functionality of the enzyme while running in the ATP hydrolysis direction. The chemical properties have been manipulated via mutagenesis and cysteine modification with methanethiosulfonate to determine which amino acid characteristics are necessary for function. Functionality is being tested using a hydrolysis based proton pumping assay as well as an ATP synthesis assay.
