Affinity modification of E1-form of Na+, K+ -ATPase revealed Asp-710 in the catalytic site

Yu A. Ovchinnikov, K. N. Dzhandzugazyan, S. V. Lutsenko, A. A. Mustayev, N. N. Modyanov

Research output: Contribution to journalArticlepeer-review

62 Scopus citations


An alkylating ATP analogue, γ-[4-(N-2-chlorethyl-N-methylamino)]benzylamide ATP (ClRATP), covalently binds to the catalytic α-subunit of Na+, K+ -ATPase yielding a product resistant to hydrolysis by the enzyme and inhibiting the ATP-hydrolysing activity. The Na+ -form of the membrane-bound Na+, K+ -ATPase modified with ClRATP was hydrolysed by pepsin under conditions providing maximum stability of the modification product (4°C, pH 1.5). The modified peptide was isolated by HPLC and its amino acid sequence was found to involve residues 706-713 of the α-subunit polypeptide chain. This fragment located near the γ-phosphate of ATP is a component of the active site. It is highly homologous with corresponding regions of the catalytic subunits of all the known E1-E2 ATPases. In the Na+ -(or E1-)enzyme form Asp-710 is the target of modification. Evidently E1- and E2-enzymes have different targets in CIRATP modification, i.e. the polypeptide chain regions near the ATP γ-phosphate in the enzyme active site differ somewhat in their conformations.

Original languageEnglish (US)
Pages (from-to)111-116
Number of pages6
JournalFEBS Letters
Issue number1
StatePublished - Jun 8 1987
Externally publishedYes


  • Affinity modification
  • Catalytic site structure
  • E-conformation
  • Na, K -ATPase

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology


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