PMID: 6411728Sep 25, 1983Paper

Antibodies to a defined region of pp60src neutralize the tyrosine-specific kinase activity.

The Journal of Biological Chemistry
L E GentryE G Krebs

Abstract

Site-specific antibodies to pp60src, the transforming protein of Rous sarcoma virus (RSV), have been prepared by immunizing rabbits with a chemically synthesized pentadecapeptide corresponding to residues 498-512 (Cys-Trp-Arg-Lys-Asp-Pro-Glu-Glu-Arg-Pro-Thr-Phe-Lys-Tyr-Leu) as deduced from the nucleotide sequence of the Prague C src gene. Antibodies specific for the synthetic peptide were purified from immune sera by affinity chromatography on peptide-bound Sepharose and characterized by a number of immunocytochemical techniques. Immunoprecipitation and Western blot analyses of normal and RSV-transformed cell lines revealed that this peptide antibody identified the authentic viral src gene product. This finding was further supported by indirect immunofluorescence on RSV-transformed rat kidney cells. The anti-peptide antibodies produced dramatic intracellular staining patterns characteristic of the src protein. Although able to immunoprecipitate pp60src, in vitro kinase reactions indicated that, unlike sera from RSV-induced tumor-bearing rabbits, the peptide antibody did not serve as a phosphate acceptor in the immunocomplex. Moreover, immunoprecipitates of pp60src prepared from this site-specific immune reagent were unable to p...Continue Reading

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