Journal Article

Fluorescein 5'-Isothiocyanate-Modified Na<sup>+</sup>, K<sup>+</sup>-ATPase, at Lys-501 of the α-Chain, Accepts ATP Independent of Pyridoxal 5'-Diphospho-5'-Adenosine Modification at Lys-480

Takeo Tsuda, Shunji Kaya, Hiroshi Funatsu, Yutaro Hayashi and Kazuya Taniguchi

in The Journal of Biochemistry

Published on behalf of The Japanese Biochemical Society

Volume 123, issue 1, pages 169-174
Published in print January 1998 | ISSN: 0021-924X
Published online January 1998 | e-ISSN: 1756-2651 | DOI: http://dx.doi.org/10.1093/oxfordjournals.jbchem.a021906
Fluorescein 5'-Isothiocyanate-Modified Na+, K+-ATPase, at Lys-501 of the α-Chain, Accepts ATP Independent of Pyridoxal 5'-Diphospho-5'-Adenosine Modification at Lys-480

Show Summary Details

Preview

The modification of Na+, K+-ATPase with increasing pyridoxal 5'-diphospho-5'-adenosine (AP2PL) concentrations resulted in saturation of the ∼0.5 mol AP2PL probe incorporation into the Lys-480/mol catalytic α-chain and reduced the Na+, K+-ATPase activity to around half without affecting the phosphorylation by acetyl phosphate (AcP), and led to increases in the AP2PL fluorescence caused by ATP and AcP. Further modification with fluorescein 5'-isothiocyanate (FITC) resulted in ∼0.9 mol FITC probe incorporation into the Lys-501/ mol α-chain and reduced the activity to below 5% without affecting the phosphorylation by AcP and these fluorescence increases. The ATP binding capacity of the AP2PL-FITC enzyme was shown to be at least 50% of that of the control enzyme (∼0.8 mol/mol α-chain). This is the first direct demonstration that Na+-bound FITC-modified enzymes accept ATP with an affinity for ATP (K1/2 > 150 μM) reduced by two orders of magnitude. The data also suggest half site reactivity of Lys-480 as to AP2PL and all site reactivity of Lys-501 as to FITC in the catalytic subunits.

Keywords: ATP binding; conformation change; fluorescein; Na−; K+-ATPase; pyridoxal

Journal Article.  0 words. 

Subjects: Biochemistry

Full text: subscription required

How to subscribe Recommend to my Librarian

Users without a subscription are not able to see the full content. Please, subscribe or login to access all content.