ATP binding site of mitochondrial creatine kinase. Affinity labelling of Asp-335 with C1RATP

Peter James, Markus Wyss, Svetlana Lutsenko, Theo Wallimann, Ernesto Carafoli

Research output: Contribution to journalArticle

Abstract

The ATP binding site of mitochondrial creatine kinase from chicken heart has been studied by modifying the purified enzyme with a 14C-labelled ATP analogue, CIRATP, in which the reactive label was covalently bound to the γ-phosphote group of ATP. The modified enzyme was digested by pepsin, and a single radioactive nonapeptide was isolated by HPLC. Amino acid analysis and direct sequence determination revealed that the isolated peptide corresponds to amino acids 335-343 within the C-terminal region of Mi-CK, this peptide being highly preserved throughout evolution. Asp-335 is very likely the site of modification by CIRATP. The specificity of the ATP analogue for the active site of creatine kinase was demonstrated by the inhibition of the enzymatic activity of Mi-CK by C1RATP and by the prevention of this inhibition bij ADP.

Original languageEnglish (US)
Pages (from-to)139-143
Number of pages5
JournalFEBS Letters
Volume273
Issue number1-2
DOIs
StatePublished - Oct 29 1990
Externally publishedYes

Fingerprint

Mitochondrial Form Creatine Kinase
Labeling
Adenosine Triphosphate
Binding Sites
Amino Acids
Peptides
Pepsin A
Enzymes
Creatine Kinase
Adenosine Diphosphate
Sequence Analysis
Labels
Chickens
Catalytic Domain
High Pressure Liquid Chromatography

Keywords

  • ATP binding site
  • CIRATP analogue
  • Creatine kinase

ASJC Scopus subject areas

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

Cite this

ATP binding site of mitochondrial creatine kinase. Affinity labelling of Asp-335 with C1RATP. / James, Peter; Wyss, Markus; Lutsenko, Svetlana; Wallimann, Theo; Carafoli, Ernesto.

In: FEBS Letters, Vol. 273, No. 1-2, 29.10.1990, p. 139-143.

Research output: Contribution to journalArticle

James, Peter ; Wyss, Markus ; Lutsenko, Svetlana ; Wallimann, Theo ; Carafoli, Ernesto. / ATP binding site of mitochondrial creatine kinase. Affinity labelling of Asp-335 with C1RATP. In: FEBS Letters. 1990 ; Vol. 273, No. 1-2. pp. 139-143.
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