Chimeras of X+,K+-ATPases - The M1-M6 region of Na+,K+-atpase is required for Na+-activated ATPase activity, whereas the M7-M10 region of H+,K+-atpase is involved in K+ de-occlusion

Citation
Jb. Koenderink et al., Chimeras of X+,K+-ATPases - The M1-M6 region of Na+,K+-atpase is required for Na+-activated ATPase activity, whereas the M7-M10 region of H+,K+-atpase is involved in K+ de-occlusion, J BIOL CHEM, 276(15), 2001, pp. 11705-11711
Citations number
48
Categorie Soggetti
Biochemistry & Biophysics
Journal title
JOURNAL OF BIOLOGICAL CHEMISTRY
ISSN journal
00219258 → ACNP
Volume
276
Issue
15
Year of publication
2001
Pages
11705 - 11711
Database
ISI
SICI code
0021-9258(20010413)276:15<11705:COX-TM>2.0.ZU;2-B
Abstract
In this study we reveal regions of Na+,K+-ATPase and H+,K+-ATPase that are involved in cation selectivity. A chimeric enzyme in which transmembrane ha irpin M5-M6 of H+,K+-ATPase was replaced by that of Na+,K+- ATPase was phos phorylated in the absence of Naf and showed no K+-dependent reactions. Next , the part originating from Na+,K+-ATPase was gradually increased in the N- terminal direction. We demonstrate that chimera HN16, containing the transm embrane segments one to six and intermediate loops of Na+,K+-ATPase, harbor s the amino acids responsible for Na+ specificity. Compared with Na+,K+-ATP ase, this chimera displayed a similar apparent Na+ affinity, a lower appare nt K+ affinity, a higher apparent ATP affinity, and a lower apparent vanada te affinity in the ATPase reaction. This indicates that the E2K form of thi s chimera is less stable than that of Na+,K+-ATPase, suggesting that it, li ke H+,K+-ATPase, de-occludes K+ ions very rapidly. Comparison of the struct ures of these chimeras with those of the parent enzymes suggests that the C -terminal 187 amino acids and the beta -subunit are involved in K+ occlusio n, Accordingly, chimera HN16 is not only a chimeric enzyme in structure, bu t also in function. On one hand it possesses the Na+-stimulated ATPase reac tion of Na+,K+-ATPase, while on the other hand it has the K+ occlusion prop erties of H+,K+-ATPase.