Creation of phosphorylation sites in proteins: Construction of a phosphorylatable human interferon α

B. L. Li, J. A. Langer, B. Schwartz, S. Pestka

Research output: Contribution to journalArticlepeer-review

62 Scopus citations

Abstract

A phosphorylation site was introduced into human interferon αA (IFN-αA) by site-specific mutation of the coding sequence. Three slightly different phosphorylation sites were created by using the predicted amino acid consensus sequences for phosphorylation by the cAMP-dependent protein kinase. The resultant modified interferons (IFN-αA-P) were expressed in Escherichia coli and purified. The purified proteins exhibit antiviral activity on bovine and human cells similar to that of the unmodified IFN-αA. The IFN-αA-P proteins can be phosphorylated by the catalytic subunit of cAMP-dependent protein kinase with [γ-32P]ATP to high specific activity (2000-5000 Ci/mmol; 1 Ci = 37 GBq) with retention of biological activity. The 32P-labeled IFN-αA-P proteins bind to cells and can be covalently bound to the IFN-α/β receptor with a bifunctional reagent as can human IFN-αA. The introduction of phosphorylation sites into proteins provides a procedure to prepare a large variety of radioactive proteins for research and clinical use.

Original languageEnglish (US)
Pages (from-to)558-562
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume86
Issue number2
DOIs
StatePublished - 1989

All Science Journal Classification (ASJC) codes

  • General

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