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Probing the topography of the intramembrane part of Na + ,K + ‐ATPase by photolabelling with 3‐(trifluoromethyl)‐3_( m [ 125 I]iodophenyl)diazirine Analysis of the hydrophobic domain of the β‐subunit
Author(s) -
Chertova E.N.,
Lutsenko S.V.,
Levi.B.,
Modyanov N.N.
Publication year - 1989
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(89)80999-8
Subject(s) - diazirine , protein subunit , chemistry , trifluoromethyl , stereochemistry , atpase , enzyme , membrane , crystallography , biochemistry , alkyl , organic chemistry , gene
The mutual disposition of the α‐helical intramembrane rods in Na + ,K + ‐ATPase subunits was studied by photolabelling of the membrane‐bound enzyme with 3‐(trifluoromethyl)‐3‐( m ‐[ 125 I]iodophenyl)diazirine ([ 125 I]TID). Under the chosen conditions for modification, the ratio of label incorporated into the subunits was found to be α/β = 2.2, demonstrating the peripheral location of the β‐subunit in the oligomeric membrane complex. The [ 125 I]TID‐labelled β‐subunit was subjected to tryptic hydrolysis and the modified fragment (Thr 27 ‐Arg 71 ) was isolated. The labelled amino acid residues are located predominantly on one side of the helix, which is helpful in unravelling the spatial orientation of the β‐subunit relative to the α‐subunit.