Abstract
An expression vector was modified to permit the rapid synthesis of purified, 32P-labeled, glutathione S-transferase (GST)-retinoblastoma (RB) fusion proteins. The products were used to screen lambda gt11 expression libraries, from which we cloned a cDNA encoding a polypeptide (RBAP-1) capable of binding directly to a putative functional domain (the pocket) of the retinoblastoma gene product (RB). The RB "pocket" is known to bind, directly or indirectly, to the cellular transcription factor, E2F, implicated in cell growth control. We have found that RBAP-1 copurifies with E2F, interacts specifically with the adenovirus E4 ORF 6/7 protein, binds specifically and directly to a known E2F DNA recognition sequence, and contains a functional tranasactivation domain. Therefore, RBAP-1 is a species of E2F and can bind specifically to the RB pocket.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Sequence
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Base Sequence
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Binding, Competitive
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Carrier Proteins / biosynthesis
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Carrier Proteins / chemistry*
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Carrier Proteins / metabolism
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Cell Cycle Proteins*
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DNA / metabolism
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DNA-Binding Proteins*
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E2F Transcription Factors
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Gene Expression Regulation
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Genetic Vectors
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Intracellular Signaling Peptides and Proteins*
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Molecular Sequence Data
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Recombinant Proteins / biosynthesis
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Retinoblastoma Protein / metabolism*
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Retinoblastoma-Binding Protein 1
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Retinoblastoma-Binding Protein 2
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Transcription Factor DP1
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Transcription Factors / chemistry*
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Transcription Factors / metabolism
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Tumor Suppressor Proteins*
Substances
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Carrier Proteins
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Cell Cycle Proteins
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DNA-Binding Proteins
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E2F Transcription Factors
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Intracellular Signaling Peptides and Proteins
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Recombinant Proteins
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Retinoblastoma Protein
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Retinoblastoma-Binding Protein 1
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Transcription Factor DP1
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Transcription Factors
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Tumor Suppressor Proteins
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DNA
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KDM5A protein, human
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Retinoblastoma-Binding Protein 2