5. The SH3 domain is about 60 amino acids long. It recognizes and binds to structural motifs in other proteins. The motif recognized by SH3 domains was found by constructing a fusion protein between an SH3 domain and glutathione-S-transferase (GST). GST fusions allow for easy purification fusing a glutathione affinity column that binds GST specifically. After tagging the purified GST-SH3 protein with biotin to make it easy to detect it was used to screen filters containing E. coli colonies expressing a cDNA library. Two different clones were found to bind to the SH3 domain: in both cases binding was shown to occur at short proline-rich sequences.
a. Could you use biotin tagged GST-SHE proteins in the same way to find cDNA domains? Why? Explain.
b. Many proteins bind to short strings of amino acids in other proteins. How do you think these kinds of interactions differ from the binds of interactions found between protein subunits of multi-subunit enzymes?
5. The SH3 domain is about 60 amino acids long. It recognizes and binds to structural motifs in other proteins. The motif recognized by SH3 domains was found by constructing a fusion protein between an SH3 domain and glutathione-S-transferase (GST). GST fusions allow for easy purification fusing a glutathione affinity column that binds GST specifically. After tagging the purified GST-SH3 protein with biotin to make it easy to detect it was used to screen filters containing E. coli colonies expressing a cDNA library. Two different clones were found to bind to the SH3 domain: in both cases binding was shown to occur at short proline-rich sequences.
a. Could you use biotin tagged GST-SHE proteins in the same way to find cDNA domains? Why? Explain.
b. Many proteins bind to short strings of amino acids in other proteins. How do you think these kinds of interactions differ from the binds of interactions found between protein subunits of multi-subunit enzymes?
