Expression of human insulin-like growth factor I in bacteria: use of optimized gene fusion vectors to facilitate protein purification.

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

Biochemistry

journal_title

Biochemistry

authors

Moks T,Abrahmsén L,Holmgren E,Bilich M,Olsson A,Uhlén M,Pohl G,Sterky C,Hultberg H,Josephson S

doi

10.1021/bi00391a005

subject

Has Abstract,Author List Incomplete

pub_date

1987-08-25 00:00:00

pages

5239-44

issue

17

eissn

0006-2960

issn

1520-4995

journal_volume

26

pub_type

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