Preparation and characterization of the recombinant selenomethionine analogue of insulin-like growth factor-I.
Insulin-like growth factor-I (IGF-I), a single-chain polypeptide consisting of 70 amino acids and 3 disulfide bridges, is a member of a class of growth factors that are involved in many proliferative and metabolic processes. To assist in solving the crystallographic three-dimensional structure, we have expressed a recombinant fusion protein precursor of IGF-I in a methionine auxotrophic strain of Escherichia coli grown in the presence of selenomethionine. An homogeneous preparation of selenomethionyl-IGF-I was then obtained by chemical cleavage of the fusion protein. The selenomethionine analogue of IGF-I was characterized by electrospray mass spectrometry, peptide mapping, analytical chromatography, and electrophoresis as well as by biological assays. The final preparation of IGF-I was found to incorporate about 90% of selenium and fully retained the functional activity.
De Bree
F M
FM
Department of Chemistry, University of York, Heslington, York, United Kingdom.
Brzozowski
A M
AM
Gellerfors
P
P
Karlsson
G
G
Rönnholm
H
H
Breme
U
U
Caccia
P
P
Taylor
G
G
Orsini
G
G
D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D007334:Insulin-Like Growth Factor I / Q000031:analogs & derivatives / Q000235:genetics* / Q000378:metabolism; D010449:Peptide Mapping; D011869:Radioligand Assay; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012645:Selenomethionine / Q000737:chemistry* / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D012645:Selenomethionine