Evaluation of the role of electrostatic residues in human epidermal growth factor by site-directed mutagenesis and chemical modification
✍ Scribed by Salil K. Niyogi; Stephen R. Campion; Douglas K. Tadaki
- Publisher
- John Wiley and Sons
- Year
- 1992
- Tongue
- English
- Weight
- 656 KB
- Volume
- 50
- Category
- Article
- ISSN
- 0730-2312
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✦ Synopsis
Four residues in the carboxy-terminal domain of human epidermal growth factor (hEGF), glutamate 40, glutamine 43, arginine 45, and aspartate 46 were targeted for site-directed mutagenesis to evaluate their potential role in epidermal growth factor (EGF) receptor-ligand interaction. One or more mutations were generated at each of these sites and the altered recombinant hEGF gene products were purified and evaluated by radioreceptor competition binding assay. Charge-conservative replacement of glutamate 40 with aspartate resulted in a decrease in receptor binding affinity to 30% relative to wild-type hEGF. On the other hand, removal of the electrostatic charge by substitution of glutamate 40 with glutamine or alanine resulted in only a slightly greater decrease in receptor binding to 25% relative receptor affinity. The introduction of a positive charge upon substitution of glutamine 43 with lysine had no effect on receptor binding. The substitution of arginine 45 with lysine also showed no effect on receptor binding, unlike the absolute requirement observed for the arginine side-chain at position 41 [Engler DA,