The protein-tyrosine kinase substrate, calpactin I heavy chain (p36), is part of the primer recognition protein complex that interacts with DNA polymerase α

H. K. Jindal, W. G. Chaney, C. W. Anderson, R. G. Davis, J. K. Vishwanatha

Research output: Contribution to journalArticle

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Abstract

Primer recognition proteins (PRP) stimulate the activity of DNA polymerase α on DNA substrates with long single-stranded template containing few primers. Purified PRP from HeLa cells and human placenta are composed of two subunits of 36,000 (PRP 1) and 41,000 (PRP 2) daltons. By amino acid sequence homology, we have identified PRP 2 as the glycolytic enzyme 3-phosphoglycerate kinase. Here we present data that establishes PRP 1 to be the protein-tyrosine kinase substrate, calpactin I heavy chain. Amino acid sequence analysis of six tryptic peptides of PRP 1 followed by homology search in a protein sequence data base revealed 100% identity of all six peptides with the deduced amino acid sequence of human calpactin I heavy chain. The activities of PRP and calpactin I coelute on gel filtration columns, and a high correlation of PRP and calpactin I activities was seen at different stages of purification. A rabbit polyclonal anti-chicken calpactin I antibody was shown to cross-react with PRP 1 polypeptide at various stages of PRP purification, and the homogeneous preparation of PRP exhibits 3-phosphoglycerate kinase (PRP 2) and calpactin I (PRP 1) activities. PRP activity is neutralized by a mouse monoclonal anti-calpactin II antibody although having no effect on the polymerase α activity itself. Calpactin II has a 50% amino acid sequence homology with calpactin I. However, PRP 1 is not calpactin II as shown by lack of cross-reaction to a monoclonal anti-calpactin II antibody on Western blots. Calpactin I and 3-phosphoglycerate kinase, purified independently, cannot be efficiently reconstituted into the PRP complex, indicating that their association in the PRP complex involves specific protein-protein interactions that remain to be elucidated. The biochemical and immunological data presented here revealing the identity of PRP 1 as calpactin I provide evidence for one physiological role of calpactin I in the cell.

Original languageEnglish (US)
Pages (from-to)5169-5176
Number of pages8
JournalJournal of Biological Chemistry
Volume266
Issue number8
StatePublished - Jul 11 1991

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Annexin A2
DNA-Directed DNA Polymerase
Protein-Tyrosine Kinases
Substrates
Proteins
Annexin A1
Phosphoglycerate Kinase
Amino Acids
Amino Acid Sequence Homology
Peptides
Purification
Antibodies

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

The protein-tyrosine kinase substrate, calpactin I heavy chain (p36), is part of the primer recognition protein complex that interacts with DNA polymerase α. / Jindal, H. K.; Chaney, W. G.; Anderson, C. W.; Davis, R. G.; Vishwanatha, J. K.

In: Journal of Biological Chemistry, Vol. 266, No. 8, 11.07.1991, p. 5169-5176.

Research output: Contribution to journalArticle

Jindal, H. K. ; Chaney, W. G. ; Anderson, C. W. ; Davis, R. G. ; Vishwanatha, J. K. / The protein-tyrosine kinase substrate, calpactin I heavy chain (p36), is part of the primer recognition protein complex that interacts with DNA polymerase α. In: Journal of Biological Chemistry. 1991 ; Vol. 266, No. 8. pp. 5169-5176.
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