Mutant epidermal growth factor polypeptides, nucleic acids, and uses therefor
View Patent ↗The present invention is based, in part, on our discovery that EGF can be engineered to generate mutants that bind to the EGF receptor (EGFR) of a cell and that have a desirable effect on the activity of the cell. For example, the mutants can agonize the receptor (i.e., increase a biological activity of the receptor), or antagonize the receptor (i.e., decrease or inhibit a biological activity of the receptor). In turn, the rate at which the cell proliferates, for example, can be changed. Moreover, some of these mutants bind EGFR with a higher affinity than wild-type EGF exhibits. The affinity may increase by about, for example, 2-, 5-, 10-, 15-, 20-, 25-, 30-, 50-, or 100-fold relative to wild-type EGF.
1. An isolated polypeptide comprising the sequence of a mutant epidermal growth factor (EGF) that conforms to the amino acid sequence X 1 -X 2 -X 3 -X 4 -X 5 -C-P-X 8 -X 9 -X 10 -X 11 -G-X 13 -C-L-X 16 -X 17 -G-X 19 -C-X 21 -Y-I-X 25 -X 26 -D-X 28 -Y-X 30 -C-X 32 -C-X 34 -X 35 -G-Y-X 38 -G-E-R-C-Q-Y-X 45 -X 46 -L-X 48 -X 49 -X 50 -X 51 -X 52 -X 53 (SEQ ID NO:47), wherein X represents any amino acid residue except cysteine (C); the sequence of the mutant EGF differs from SEQ ID NO:1 (wild type EGF) by including 3-10 amino acid substitutions; and the mutant EGF comprises amino acid substitutions at X 48 , X 51 , and X 52 .
2. The isolated polypeptide of claim 1 , wherein the mutant EGF comprises 3, 4, 5, 6, 7, or 8 amino acid substitutions.
3. The isolated polypeptide of claim 1 , wherein X 48 is R or T.
4. The isolated polypeptide of claim 1 , wherein X 51 is G, A, W, K, or Y.
5. The isolated polypeptide of claim 1 , wherein X 52 is P, R, or T.
6. The isolated polypeptide of claim 1 , wherein the mutant EGF consists of SEQ ID NO:3, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:36, or SEQ ID NO:38.
7. The isolated polypeptide of claim 1 , wherein the polypeptide consists of the mutant EGF.
8. The isolated polypeptide of claim 6 , wherein the polypeptide consists of the mutant EGF.
9. The isolated polypeptide of claim 1 , wherein the mutant EGF binds a human EGF receptor.
10. The isolated polypeptide of claim 6 , wherein the mutant EGF binds a human EGF receptor with at least five-fold higher affinity than SEQ ID NO:1 (wild type EGF) binds the EGF receptor.
11. A kit comprising: a polypeptide of claim 1 and instructions for diagnostic or therapeutic use.