4HYT

Na,K-ATPase in the E2P state with bound ouabain and Mg2+ in the cation-binding site

Summary for 4HYT

• Entry DOI: 10.2210/pdb4hyt/pdb
• Related: 3KDP 3N23
• Descriptor: Sodium/potassium-transporting ATPase subunit alpha-1, O-[(S)-({(2R)-2,3-bis[(9Z)-octadec-9-enoyloxy]propyl}oxy)(hydroxy)phosphoryl]-L-serine, O-DODECANYL OCTAETHYLENE GLYCOL, ... (13 entities in total)
• Functional Keywords: membrane transporter, hydrolase-membrane protein complex, hydrolase/membrane protein
• Biological source: Sus scrofa (pig); More
• Total number of polymer chains: 6
• Total formula weight: 320755.17

Authors

Laursen, M.,Yatime, L.,Nissen, P.,Fedosova, N.U. (deposition date: 2012-11-14, release date: 2013-06-26, Last modification date: 2023-09-20)

Primary citation

Laursen, M.,Yatime, L.,Nissen, P.,Fedosova, N.U.
Crystal structure of the high-affinity Na+,K+-ATPase-ouabain complex with Mg2+ bound in the cation binding site.
Proc.Natl.Acad.Sci.USA, 110:10958-10963, 2013

PubMed Abstract: The Na(+),K(+)-ATPase maintains electrochemical gradients for Na(+) and K(+) that are critical for animal cells. Cardiotonic steroids (CTSs), widely used in the clinic and recently assigned a role as endogenous regulators of intracellular processes, are highly specific inhibitors of the Na(+),K(+)-ATPase. Here we describe a crystal structure of the phosphorylated pig kidney Na(+),K(+)-ATPase in complex with the CTS representative ouabain, extending to 3.4 Å resolution. The structure provides key details on CTS binding, revealing an extensive hydrogen bonding network formed by the β-surface of the steroid core of ouabain and the side chains of αM1, αM2, and αM6. Furthermore, the structure reveals that cation transport site II is occupied by Mg(2+), and crystallographic studies indicate that Rb(+) and Mn(2+), but not Na(+), bind to this site. Comparison with the low-affinity [K2]E2-MgF(x)-ouabain structure [Ogawa et al. (2009) Proc Natl Acad Sci USA 106(33):13742-13747) shows that the CTS binding pocket of [Mg]E2P allows deep ouabain binding with possible long-range interactions between its polarized five-membered lactone ring and the Mg(2+). K(+) binding at the same site unwinds a turn of αM4, dragging residues Ile318-Val325 toward the cation site and thereby hindering deep ouabain binding. Thus, the structural data establish a basis for the interpretation of the biochemical evidence pointing at direct K(+)-Mg(2+) competition and explain the well-known antagonistic effect of K(+) on CTS binding.

PubMed: 23776223
DOI: 10.1073/pnas.1222308110

Experimental method

X-RAY DIFFRACTION (3.404 Å)