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. 1989 Jan;86(2):558–562. doi: 10.1073/pnas.86.2.558

Creation of phosphorylation sites in proteins: construction of a phosphorylatable human interferon alpha.

B L Li 1, J A Langer 1, B Schwartz 1, S Pestka 1
PMCID: PMC286511  PMID: 2911594

Abstract

A phosphorylation site was introduced into human interferon alpha A (IFN-alpha A) 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-alpha A-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-alpha A. The IFN-alpha A-P proteins can be phosphorylated by the catalytic subunit of cAMP-dependent protein kinase with [gamma-32P]ATP to high specific activity (2000-5000 Ci/mmol; 1 Ci = 37 GBq) with retention of biological activity. The 32P-labeled IFN-alpha A-P proteins bind to cells and can be covalently bound to the IFN-alpha/beta receptor with a bifunctional reagent as can human IFN-alpha A. The introduction of phosphorylation sites into proteins provides a procedure to prepare a large variety of radioactive proteins for research and clinical use.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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