Hot and cold running ion pumps

S.L. Myers,¹ F. Cornelius,² H.-J. Apell³ and R.J. Clarke,¹ ¹School of Chemistry, University of Sydney, NSW 2006, Australia, ²Department of Physiology and Biophysics, University of Aarhus, DK-8000 Aarhus C, Denmark and ³Faculty of Biology, University of Constance, D-78435 Constance, Germany.

Crystal structures of the Na⁺,K⁺-ATPase from both a warm-blooded animal (pig) (Morth et al., 2007) and a cold-blooded animal (shark) (Shinoda et al., 2009) have recently been published. Although the structures of the enzyme from these two species appear very similar, we have discovered major differences in their kinetics,

From investigations of K⁺ occlusion by the phosphoenzyme intermediate of the Na⁺,K⁺-ATPase and its K⁺-stimulated dephosphorylation via stopped-flow fluorimetry we have found that, whereas both enzymes appear to have similar rate constants of K⁺-occlusion of 370-380 s^-1, the two enzymes have very different rate constants of dephosphorylation. For the shark enzyme, dephosphorylation proceeds with a rate constant of only 48 (±2) s^-1 at 24°C and pH 7.4, whereas for the pig enzyme the rate constant is >365 s^-1. The dephosphorylation is, thus, the major rate-determining step of the shark enzyme under saturating concentrations of all substrates. For the pig enzyme, on the other hand, the major rate-determining step under the same conditions is the conformational E2-E1 transition of unphosphorylated enzyme and its associated K⁺ release to the cytoplasm. The differences in rate constant of the dephosphorylation reaction of the two enzymes are paralleled by compensating changes to the rate constant for the E2-E1 transition (Kahlid et al., 2010), which explains why the differences in the enzymes' kinetic behaviour have not previously been identified in steady-state kinetic studies of the enzyme's entire reaction cycle.

In mammals, heat generation by the Na⁺,K⁺-ATPase as a by-product of ion pumping is thought to make approximately a 12% contribution towards the maintenance of body temperature. Therefore, the possibility should be investigated whether under physiological conditions the differences in Na⁺,K⁺-ATPase kinetics, which we have identified between a warm- and a cold-blooded animal, could in part be responsible for the higher body temperature of warm-blooded animals.

Khalid M, Cornelius F, Clarke RJ. (2010) Biophysical Journal 98: 2290.

Morth JP, Pedersen BP, Toustrup-Jensen MS, Sørensen TL, Petersen J, Andersen JP, Vilsen B, Nissen P. (2007) Nature 450: 1043-9.

Shinoda T, Ogawa H, Cornelius F, Toyoshima C (2009) Nature 459: 446-50.