Evidence for the ability of nucleophosmin/B23 to bind ATP

Jei H. Chang, Jui Y. Lin, Ming H. Wu, Yat Ming Yung

Research output: Journal article publicationJournal articleAcademic researchpeer-review

22 Citations (Scopus)

Abstract

By taking advantage of its ability to be retained by ATP-agarose, we have demonstrated that nucleophosmin/B23 is capable of binding ATP. The specificity of the binding was confirmed by the absence of significant binding to AMP-agarose and by its loss when nucleophosmin/B23 in nuclear extracts was preincubated with ATP. Preincubation of the nuclear extracts with other ribonucleotide triphosphates (GTP, CTP, UTP) did not compete for the binding of nucleophosmin/B23 to ATP-agarose. The purified recombinant nucleophosmin/B23 was also able to be retained by ATP-agarose. The K(d) for binding of ATP to the purified recombinant nucleophosmin/B23, on the basis of retention on a nitrocellulose membrane, was 86.5 ± 8.3 μM; the number of binding sites was 0.68 per nucleophosmin/B23 protein molecule. To determine the possible ATP-binding site of nucleophosmin/B23, various deletion clones including the two mutants in which the putative ATP-binding sequence had been deleted were constructed. Deletion of the portions of the molecule (residues 83-152 and 185-240) had little effect on the ATP binding. The C-terminal deleted mutant (residue 242 to the C-terminus deleted) lost most of its ability to be retained by ATP-agarose and to bind [α-32P]ATP on a nitrocellulose membrane. The results indicate that the C-terminal portion (residues 242-294) contains the essential ATP-binding site of nucleophosmin/B23.
Original languageEnglish
Pages (from-to)539-544
Number of pages6
JournalBiochemical Journal
Volume329
Issue number3
DOIs
Publication statusPublished - 1 Feb 1998
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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