Abstract
Full-length and N-terminal deletion mutants of human c-raf-1 cDNA were cloned into Escherichia coli expression plasmids. Bacterially expressed c-raf proteins were purified by anion-exchange, gel filtration, and affinity chromatography. Microinjection of mutant c-raf proteins into G0-arrested NIH 3T3 cells induced DNA synthesis and morphological transformation, whereas microinjection of full-length c-raf had no effect. The amino terminus of the raf protein has an important negative regulatory influence; alteration of this region resulted in increased kinase activity and oncogenicity.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Cell Transformation, Neoplastic*
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Cells, Cultured
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Cloning, Molecular
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DNA Replication*
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Escherichia coli / genetics
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Humans
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Mice
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Microinjections
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Mutation
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Protein-Tyrosine Kinases / metabolism
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Proto-Oncogene Proteins / administration & dosage
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / metabolism*
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Proto-Oncogene Proteins c-raf
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Recombinant Proteins / administration & dosage
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Recombinant Proteins / metabolism
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Restriction Mapping
Substances
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Proto-Oncogene Proteins
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Recombinant Proteins
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Protein-Tyrosine Kinases
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Proto-Oncogene Proteins c-raf