Molecular modulation of the alpha‐subunit VISIT‐DG sequence in the catalytic sites of Escherichia coli ATP synthase

Abstract

F1Fo‐ATP synthase is the smallest known biological nanomotor, found from bacteria to man. This enzyme is responsible for ATP synthesis by oxidative or photophosphorylation in membranes of bacteria, mitochondria, and chloroplasts. Thus, ATP synthase is the fundamental means of cell energy production in animals, plants, and almost all microorganisms. A typical 70 kg human with a relatively sedentary lifestyle will generate around 2.0 million kg of ATP from ADP and Pi (inorganic phosphate) in a 75‐year lifespan. Pi binding is a primary step in ATP synthesis by ATP synthase, so understanding the molecular basis of Pi binding is an important goal. X‐ray structures of the catalytic sites show the highly conserved α‐subunit VISIT‐DG sequence in close proximity to the conserved phosphate binding residues αR376, βR182, βK155, and βR246 in the phosphate binding subdomain. Lately, we found that αSer‐347 of VISIT‐DG sequence is an additional, important residue involved in phosphate‐binding and transition state stabilization. In contrast, αGly‐351, although strongly conserved and clearly important for function, appears not to play a direct role. Further, through mutagenic modulation and biochemical characterization of αVal‐345, αIle‐346, αIle‐348, αThr‐349, and αAsp‐350, we have been able to identify their direct/indirect role in Pi binding, in maintaining structural integrity of the Pi binding pocket, and in function.