Abstract
By using in vitro binding assays and the yeast two-hybrid system, we have found that a full-length rat Cdc37-related protein (RCdc37) could associate specifically with the retinoblastoma susceptibility gene product (pRB). A series of GST-RCdc37 deletion mutants was constructed to define the amino acid sequence required for the interaction with pRB. A GST-RCdc37 (-20 approximately 229) possessed an activity to associate with pRB, whereas GST-RCdc37 (-20 approximately 165) lost its activity, indicating that the amino acid sequence between 166 and 229 of RCdc37 was essential for the association with pRB. Interestingly, there exists a highly conserved pRB-binding motif (LXCXE; X = any amino acid) that is essential for pRB binding of SV40 large T antigen, E1A, and E7 proteins. A similar experiment using a pRB deletion mutant revealed that the carboxy-terminal portion of pRB was required for binding to RCdc37.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Animals
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Binding Sites
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Carrier Proteins / biosynthesis
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Carrier Proteins / isolation & purification
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Carrier Proteins / metabolism*
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Cell Cycle Proteins / biosynthesis
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Cell Cycle Proteins / isolation & purification
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Cell Cycle Proteins / metabolism*
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Conserved Sequence
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Drosophila Proteins*
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Escherichia coli
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Glutathione Transferase / biosynthesis
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Molecular Chaperones*
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Mutagenesis, Site-Directed
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Protein Biosynthesis
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Rats
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Recombinant Fusion Proteins / biosynthesis
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Recombinant Fusion Proteins / isolation & purification
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Recombinant Proteins / metabolism
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Retinoblastoma Protein / metabolism*
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Saccharomyces cerevisiae
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Saccharomyces cerevisiae Proteins
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Sequence Deletion
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Transcription, Genetic
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beta-Galactosidase / biosynthesis
Substances
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CDC37 protein, S cerevisiae
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Carrier Proteins
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Cdc37 protein, rat
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Cell Cycle Proteins
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Drosophila Proteins
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Molecular Chaperones
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Recombinant Fusion Proteins
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Recombinant Proteins
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Retinoblastoma Protein
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Saccharomyces cerevisiae Proteins
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Glutathione Transferase
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beta-Galactosidase