Abstract:
:Several fusions between the gene for human insulin-like growth factor I (IGF-I) and the genes for different IgG-binding fragments of staphylococcal protein A were assembled and compared regarding expression, secretion, and purification of the peptide hormone. After IgG affinity purification of the fusion proteins from the growth medium of Staphylococcus aureus or Escherichia coli, native IGF-I was released by cleavage of an Asn-Gly peptide bond with hydroxylamine. An optimized expression system based on a modified synthetic IgG-binding domain (z), resistant to hydroxylamine, gave the highest yield of fusion protein. After cleavage, the hormone could be separated from the IgG-binding moiety and from noncleaved fusion protein by a second passage through the IgG affinity column. The biological activity and the purity of the IGF-I obtained were confirmed by a radioreceptor assay, N-terminal sequence analysis, polyacrylamide gel electrophoresis, isoelectric focusing, and high-performance liquid chromatography.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Moks T,Abrahmsén L,Holmgren E,Bilich M,Olsson A,Uhlén M,Pohl G,Sterky C,Hultberg H,Josephson Sdoi
10.1021/bi00391a005subject
Has Abstract,Author List Incompletepub_date
1987-08-25 00:00:00pages
5239-44issue
17eissn
0006-2960issn
1520-4995journal_volume
26pub_type
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