8K1L

Cryo-EM structure of Na+,K+-ATPase alpha2 from Artemia salina in cation-free E2P form

Summary for 8K1L

• Entry DOI: 10.2210/pdb8k1l/pdb
• EMDB information: 36794
• Descriptor: Na+,K+-ATPase alpha2KK, Na+,K+-ATPase beta2, TETRAFLUOROALUMINATE ION (3 entities in total)
• Functional Keywords: p-type atpase, sodium pump, membrane protein, transporter, transport protein
• Biological source: Artemia salina; More
• Total number of polymer chains: 2
• Total formula weight: 149743.61

Authors

Abe, K.,Artigas, P. (deposition date: 2023-07-11, release date: 2023-11-29, Last modification date: 2024-10-16)

Primary citation

Artigas, P.,Meyer, D.J.,Young, V.C.,Spontarelli, K.,Eastman, J.,Strandquist, E.,Rui, H.,Roux, B.,Birk, M.A.,Nakanishi, H.,Abe, K.,Gatto, C.
A Na pump with reduced stoichiometry is up-regulated by brine shrimp in extreme salinities.
Proc.Natl.Acad.Sci.USA, 120:e2313999120-e2313999120, 2023

PubMed Abstract: Brine shrimp () are the only animals to thrive at sodium concentrations above 4 M. Salt excretion is powered by the Na,K-ATPase (NKA), a heterodimeric (αβ) pump that usually exports 3Na in exchange for 2 K per hydrolyzed ATP. express several NKA catalytic α-subunit subtypes. High-salinity adaptation increases abundance of α2, an isoform that contains two lysines (Lys308 and Lys758 in transmembrane segments TM4 and TM5, respectively) at positions where canonical NKAs have asparagines ( α1's Asn333 and Asn785). Using de novo transcriptome assembly and qPCR, we found that express two salinity-independent canonical α subunits (α1 and α3), as well as two β variants, in addition to the salinity-controlled α2. These β subunits permitted heterologous expression of the α2 pump and determination of its CryoEM structure in a closed, ion-free conformation, showing Lys758 residing within the ion-binding cavity. We used electrophysiology to characterize the function of α2 pumps and compared it to that of α1 (and its α2-mimicking single- and double-lysine substitutions). The double substitution N333K/N785K confers α2-like characteristics to α1, and mutant cycle analysis reveals energetic coupling between these two residues, illustrating how α2's Lys308 helps to maintain high affinity for external K when Lys758 occupies an ion-binding site. By measuring uptake under voltage clamp of the K-congener Rb, we prove that double-lysine-substituted pumps transport 2Na and 1 K per catalytic cycle. Our results show how the two lysines contribute to generate a pump with reduced stoichiometry allowing to maintain steeper Na gradients in hypersaline environments.

PubMed: 38079564
DOI: 10.1073/pnas.2313999120

Experimental method

ELECTRON MICROSCOPY (3.44 Å)