%0 Journal Article %A Kenta Yamamoto %A Vikas Dubey %A Katsumasa Irie %A Hanayo Nakanishi %A Himanshu Khandelia %A Yoshinori Fujiyoshi %A Kazuhiro Abe %T A single K+-binding site in the crystal structure of the gastric proton pump %D 2019 %R 10.1101/608851 %J bioRxiv %P 608851 %X The gastric proton pump (H+,K+-ATPase), a P-type ATPase responsible for gastric acidification, mediates electro-neutral exchange of H+ and K+ coupled with ATP hydrolysis, but with an as yet undetermined transport stoichiometry. Here we show crystal structures at a resolution of 2.5 Å of the pump in the E2-P transition state, in which the counter-transporting cation is occluded. We found a single K+ bound to the cation-binding site of H+,K+-ATPase, indicating an exchange of 1H+/1K+ per hydrolysis of one ATP molecule. This fulfils the energy requirement for the generation of a six pH unit gradient across the membrane. The structural basis of K+recognition is resolved, supported by molecular dynamics simulations, and this establishes how H+,K+-ATPase overcomes the energetic challenge to generate an H+ gradient of more than a million-fold – the highest cation gradient known in any mammalian tissue – across the membrane. %U https://www.biorxiv.org/content/biorxiv/early/2019/04/15/608851.full.pdf