| Autor: |
Suh SS; Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena 91125., Haymore BL, Arnold FH |
| Jazyk: |
angličtina |
| Zdroj: |
Protein engineering [Protein Eng] 1991 Feb; Vol. 4 (3), pp. 301-5. |
| DOI: |
10.1093/protein/4.3.301 |
| Abstrakt: |
Variants of bovine somatotropin have been engineered to contain synthetic metal-binding sites consisting of two solvent-exposed histidines separated by a single turn of an alpha-helix (His-X3-His variants). The affinities of these proteins for Cu(II) were characterized by measuring their partition coefficients in an aqueous two-phase polymer system. The partition coefficients were used to generate binding constants for formation of a complex between the engineered metal-binding site and Cu(II) chelated to an iminodiacetic acid derivative of polyethylene glycol. For three His-X3-His variants described here, these constants range from 2 x 10(4) to 1.6 x 10(6) M-1. The metal affinity of a His-X3-His site depends on the rigidity of the helix into which the site is engineered. The affinities of the His-X3-His sites for Cu(II) are large enough to dramatically increase not only the partitioning of these proteins in aqueous two-phase systems, but also their retention times on a metal-affinity chromatography column. Both these features can greatly facilitate the purification of engineered proteins. Criteria for choosing positions for incorporating metal-binding sites are discussed. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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