Design and Construction of Synthetic Metal-Binding Proteins
Annual rept. 1 Jul 1988-30 Jun 1989
CALIFORNIA INST OF TECH PASADENA
Pagination or Media Count:
The goal of this project is to assemble a systematic framework for design and construction of synthetic metal-binding proteins for metal recognition and recovery. In the first phase of this work, we are investigating metal-binding sites that can be built into the surface of a protein with relatively few amino acid substitutions. One such site, consisting of two histidines positioned His-X-X-X-His in an alpha-helix, has been built into yeast iso-1-cytochrome c. The resulting mutant protein binds CuII and other metals with high affinity.In addition to the protein engineering work, a novel technique for quantifying metal-protein interactions has been developed. This technique is based on the partitioning of metal-binding proteins between two aqueous-polymer phases. When polymer in one phase is derivatized with the appropriate metal chelate e.g. polyethylene glycol-IDA-CuII, partitioning of a protein between the two phases depends on the number of binding sites and the affinity of each site for the metal. We have demonstrated that metal-protein binding constants can be obtained for binding sites accessible to the chelated metals. Keywords Protein-metal interaction Binding sites.
- Genetic Engineering and Molecular Biology