TY - JOUR

T1 - The role of Glu327 of the a-subunit in coordinating Na + and K+ in binding and occlusion in Na,K-ATPase

AU - Rasmussen, J. H.

AU - Pedersen, P. A.

AU - Xielsen, J. M.

AU - Lanfermeijer, F. C.

AU - Jorgcnsen, P. L.

PY - 1997/12/1

Y1 - 1997/12/1

N2 - MutagPDPsis of Ghr2' in the 4th transmembrane segment of the rt-subuiiil of Na.K ATPase was performed to distinguish effects of mutations on the V Y.-i conformational transitions from direct interference with cation binding and nrrluriioTi. Substitutions of GIu327 for Gin reduces Na,K-ATPase activity and Na dependent phosphorylation to about 1/3 and these activities are eliminated in Glu327 Asp. The mutations do not alter high affinity binding of pH] AT P or pH]-ouabain and chyrnotryptic digestion yields normal cleavage prodiu.ts a.s an indication that the structure of rytoplasmic and extracelluiar ciornaina are preserved. The mutations abolish high affinity occlusion of 204T1+ ions and the affinities of K +-ions for displacement of [3H]-ATP arc reduced in parallel. Neither Na+ nor K+ (10 mM) are able to alter the conformationai -tate of the o subunit of Glu327Gln as monitored with chymolrypsin. while transition to the E|-form or £2-form are observed after binding of AT P 01 ouabain. Interference with occiusion of K+ in the F-f-Kj-form and of Na" in the- KiP[3Naj-form, may therefore bo due to removal of groups coordinating K+ or Na+ in the occiusion cavities.

AB - MutagPDPsis of Ghr2' in the 4th transmembrane segment of the rt-subuiiil of Na.K ATPase was performed to distinguish effects of mutations on the V Y.-i conformational transitions from direct interference with cation binding and nrrluriioTi. Substitutions of GIu327 for Gin reduces Na,K-ATPase activity and Na dependent phosphorylation to about 1/3 and these activities are eliminated in Glu327 Asp. The mutations do not alter high affinity binding of pH] AT P or pH]-ouabain and chyrnotryptic digestion yields normal cleavage prodiu.ts a.s an indication that the structure of rytoplasmic and extracelluiar ciornaina are preserved. The mutations abolish high affinity occlusion of 204T1+ ions and the affinities of K +-ions for displacement of [3H]-ATP arc reduced in parallel. Neither Na+ nor K+ (10 mM) are able to alter the conformationai -tate of the o subunit of Glu327Gln as monitored with chymolrypsin. while transition to the E|-form or £2-form are observed after binding of AT P 01 ouabain. Interference with occiusion of K+ in the F-f-Kj-form and of Na" in the- KiP[3Naj-form, may therefore bo due to removal of groups coordinating K+ or Na+ in the occiusion cavities.

UR - http://www.scopus.com/inward/record.url?scp=33750188732&partnerID=8YFLogxK

M3 - Journal article

AN - SCOPUS:33750188732

VL - 11

JO - F A S E B Journal

JF - F A S E B Journal

SN - 0892-6638

IS - 9

ER -