Differential photoaffinity labeling of catalytic subunits of NaK-ATPase with carrier-free /sup 125/I-cardiac glycosides
Abstract
The authors have obtained evidence for structural differences in the cardiac glycoside binding site between the ..cap alpha.. and ..cap alpha..(+) forms of the catalytic subunit of NaK-ATPase, using three closely related photoaffinity derivatives of the cardiotonic steroid, digitoxigenin. (/sup 125/I)N-(p-azido-m-iodo-o-hydroxybenzoyl)-4-amino-4,6-dideoxy-galactosyl digitoxigenin (IA-GaD), (/sup 125/I)N-(3-(p-azido-m-iodophenyl)-propionyl)-4-amino-4,6-dideoxy-ga-lactosyl digitoxigenin (AIPP-GaD) and (/sup 125/I)N-(3-(p-azido-m-iodophenyl)-propionyl)-4-amino-4,6-dideoxy-glucosyl digitoxi-genin (AIPP-GluD) were synthesized. AIPP-GaD and AIPP-GluD are stereoisomers. Eel electroplax and dog kidney NaK-ATPase (..cap alpha.. form) and rat brain synaptosomes (rich in ..cap alpha..(+) form) were photolabelled and then analyzed by SDS-PAGE and autoradiography. Photolysis with either carrier-free IA-GaD or AIPP-GluD gave ouabain-protectable labelling of NaK-ATPase catalytic subunit from all three tissues. However, photolysis with AIPP-GaD showed protectable labelling of the enzyme from eel and kidney but not from brain. This suggests a structural difference in the ..cap alpha..(+) form which results in either an inability to bind AIPP-GaD, or, perhaps more likely, an absence of a photoinsertion site in the correct location in the ..cap alpha..(+) form, as compared with the ..cap alpha.. form. It is of interest that the labelling pattern of the enzyme in the human erythrocyte resembles that of the brain enzyme.
- Authors:
- Publication Date:
- Research Org.:
- Univ. of Wisconsin, Madison
- OSTI Identifier:
- 6943259
- Report Number(s):
- CONF-8606151-
Journal ID: CODEN: FEPRA; TRN: 87-006998
- Resource Type:
- Conference
- Journal Name:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)
- Additional Journal Information:
- Journal Volume: 45:6; Conference: 76. annual meeting of the Federation of American Society for Experimental Biology, Washington, DC, USA, 8 Jun 1986
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; ATP-ASE; BIOLOGICAL VARIABILITY; MOLECULAR STRUCTURE; DIGITOXIN; BIOCHEMICAL REACTION KINETICS; BRAIN; DOGS; ELECTROPHORESIS; HEART; IODINE 125; ISOENZYMES; NERVES; RATS; RECEPTORS; STEROIDS; ACID ANHYDRASES; ANIMALS; BETA DECAY RADIOISOTOPES; BODY; CARBOHYDRATES; CARDIAC GLYCOSIDES; CARDIOTONICS; CARDIOVASCULAR AGENTS; CARDIOVASCULAR SYSTEM; CENTRAL NERVOUS SYSTEM; DAYS LIVING RADIOISOTOPES; DIGITALIS GLYCOSIDES; DRUGS; ELECTRON CAPTURE RADIOISOTOPES; ENZYMES; GLYCOSIDES; HYDROLASES; INTERMEDIATE MASS NUCLEI; IODINE ISOTOPES; ISOTOPES; KINETICS; MAMMALS; MEMBRANE PROTEINS; NERVOUS SYSTEM; NUCLEI; ODD-EVEN NUCLEI; ORGANIC COMPOUNDS; ORGANS; PHOSPHOHYDROLASES; PROTEINS; RADIOISOTOPES; REACTION KINETICS; RODENTS; VERTEBRATES; 550201* - Biochemistry- Tracer Techniques
Citation Formats
Lowndes, J, Hokin-Neaverson, M, and Ruoho, A. Differential photoaffinity labeling of catalytic subunits of NaK-ATPase with carrier-free /sup 125/I-cardiac glycosides. United States: N. p., 1986.
Web.
Lowndes, J, Hokin-Neaverson, M, & Ruoho, A. Differential photoaffinity labeling of catalytic subunits of NaK-ATPase with carrier-free /sup 125/I-cardiac glycosides. United States.
Lowndes, J, Hokin-Neaverson, M, and Ruoho, A. 1986.
"Differential photoaffinity labeling of catalytic subunits of NaK-ATPase with carrier-free /sup 125/I-cardiac glycosides". United States.
@article{osti_6943259,
title = {Differential photoaffinity labeling of catalytic subunits of NaK-ATPase with carrier-free /sup 125/I-cardiac glycosides},
author = {Lowndes, J and Hokin-Neaverson, M and Ruoho, A},
abstractNote = {The authors have obtained evidence for structural differences in the cardiac glycoside binding site between the ..cap alpha.. and ..cap alpha..(+) forms of the catalytic subunit of NaK-ATPase, using three closely related photoaffinity derivatives of the cardiotonic steroid, digitoxigenin. (/sup 125/I)N-(p-azido-m-iodo-o-hydroxybenzoyl)-4-amino-4,6-dideoxy-galactosyl digitoxigenin (IA-GaD), (/sup 125/I)N-(3-(p-azido-m-iodophenyl)-propionyl)-4-amino-4,6-dideoxy-ga-lactosyl digitoxigenin (AIPP-GaD) and (/sup 125/I)N-(3-(p-azido-m-iodophenyl)-propionyl)-4-amino-4,6-dideoxy-glucosyl digitoxi-genin (AIPP-GluD) were synthesized. AIPP-GaD and AIPP-GluD are stereoisomers. Eel electroplax and dog kidney NaK-ATPase (..cap alpha.. form) and rat brain synaptosomes (rich in ..cap alpha..(+) form) were photolabelled and then analyzed by SDS-PAGE and autoradiography. Photolysis with either carrier-free IA-GaD or AIPP-GluD gave ouabain-protectable labelling of NaK-ATPase catalytic subunit from all three tissues. However, photolysis with AIPP-GaD showed protectable labelling of the enzyme from eel and kidney but not from brain. This suggests a structural difference in the ..cap alpha..(+) form which results in either an inability to bind AIPP-GaD, or, perhaps more likely, an absence of a photoinsertion site in the correct location in the ..cap alpha..(+) form, as compared with the ..cap alpha.. form. It is of interest that the labelling pattern of the enzyme in the human erythrocyte resembles that of the brain enzyme.},
doi = {},
url = {https://www.osti.gov/biblio/6943259},
journal = {Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)},
number = ,
volume = 45:6,
place = {United States},
year = {Thu May 01 00:00:00 EDT 1986},
month = {Thu May 01 00:00:00 EDT 1986}
}