Crystallization of IGF-1
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Abstract
Crystalline IGF-1 is provided along with a method for production thereof. Crystallizing IGF-1 comprises the steps of mixing an aqueous solution comprising IGF-1 with a reservoir solution comprising a precipitant to form a mixture; and crystallizing the mixture, optionally also recrystallizing and isolating the crystalline IGF-1. In addition, a method for identifying IGF-1 indirect agonists is provided using a detergent as a standard for the level of inhibition of binding of IGFBP-1 or IGFBP-3 to IGF-1 and/or using the coordinates of the binding pockets of IGF-1 to which a candidate indirect agonist binds for structure-based drug design.
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Citations
35 Claims
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1-27. -27. (canceled)
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28. A method of identifying indirect agonists of IGF-1 comprising the steps of:
- (a) comparing the ability of N,N-bis(3-D-gluconamidopropyl)-deo-xycholamine to inhibit binding of IGFBP-1 or -3 to IGF-1 with the ability of a candidate indirect agonist of IGF-1 to so inhibit binding; and
(b) determining whether the candidate agonist inhibits such binding at least as well as N,N-bis(3-D-gluconamidopropyl)-deoxycholamine. - View Dependent Claims (29, 30)
- (a) comparing the ability of N,N-bis(3-D-gluconamidopropyl)-deo-xycholamine to inhibit binding of IGFBP-1 or -3 to IGF-1 with the ability of a candidate indirect agonist of IGF-1 to so inhibit binding; and
- 31. A method of identifying indirect agonists of IGF-1 comprising co-crystallizing a candidate indirect agonist of IGF-1 with IGF-1 to form a co-crystalline structure and determining if the candidate agonist binds to one or both of two patches on IGF-1, wherein one patch has the amino acid residues Glu 3, Thr 4, Leu 5, Asp 12, Ala 13, Phe 16, Val 17, Cys 47, Ser 51, Cys 52, Asp 53, Leu 54, and Leu 57, and the second patch has the amino acid residues Val 11, Gln 15, Phe 23, Phe 25, Asn 26, Val 44, Phe 49, and Arg 55, and wherein binding occurs if there is at least one contact between each listed amino acid residue of a given patch and the candidate agonist that is less than or equal to 6 angstroms in the co-crystalline structure.
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35-50. -50. (canceled)
Specification