Simultaneous isolation of insulin-like growth factors I and II from adult sheep serum.
Ovine insulin-like growth factors I and II (oIGF-I and oIGF-II) have been purified from adult sheep serum. oIGF-II-like receptor-binding activity and IGF-I-like immunoactivity were enriched on SP-Sephadex C-25, then purified using HPLC in the presence of a variety of counter ions. IGF-I- and IGF-II-like activities were separated using HPLC in the presence of 0.2% tetrabutylammonium phosphate at pH 7.0. The final recovery of oIGF-I was 82.6 micrograms from 3.2 litres of adult sheep serum (a yield of 17.6%), and the recovery of oIGF-II was 388 micrograms (a yield of 13.3%). Both IGF preparations were considered to be homogeneous as judged by single sharp peaks during analytical HPLC, and unique N-terminal amino acid sequences. Purified ovine IGFs had molecular weights similar to that of other IGFs (approximately 7000), and the first 30 N-terminal amino acids of both peptides were identical to their human counterparts. The isoelectric points of oIGF-I (pI approximately 8.2) and oIGF-II (pI approximately 6.8) were similar to those of human (h) IGFs (hIGF-I pI approximately 8.2; hIGF-II pI approximately 6.5), and the overall amino acid content of the ovine IGFs was also similar to that of IGFs from other species. oIGF-II preparations from fetal sheep and from adult sheep appeared to be identical. The isolation procedure represents one of general utility that can be easily modified to facilitate the isolation of recombinant IGFs from culture fluid.
Hey
A W
AW
Department of Physiology, Monash University, Clayton, Victoria, Australia.
Browne
C A
CA
Simpson
R J
RJ
Thorburn
G D
GD
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000097:blood / Q000302:isolation & purification*; D007335:Insulin-Like Growth Factor II / Q000097:blood / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D012756:Sheep; D013002:Somatomedins / Q000302:isolation & purification*
D000596:Amino Acids; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II