Structure-function studies of an IGF-I analogue that can be chemically cleaved to a two-chain mini-IGF-I.
The structure and biological activities of two disulphide isomers of a C-region deletion mutant of insulin-like growth factor-I (IGF-I) which has an Asn--Gly link engineered at the junction of the A- and B-regions were studied before and after chemical cleavage. Circular dichroism (CD) spectra and binding affinity to IGF binding protein 3 (IGFBP3) indicated that the treatment with hydroxylamine did not disrupt the overall tertiary fold of the hormones. Cleavage restored some binding affinity for the IGF-I receptor in both isomers and weakly restored the ability to stimulate incorporation of tritiated thymidine into DNA in NIH 3T3 fibroblasts transfected with the human IGF-I receptor. Cleavage also restored metabolic capacity, as measured by the ability of the isomers to promote lipogenesis in isolated rat adipocytes through the insulin receptor. These results are consistent with the theory that binding of IGF-I to the IGF-I receptor requires a conformational change similar to that involved in insulin binding the insulin receptor. The weak affinity for the IGF-I receptor after cleavage is consistent with the belief that residues in the C-region interact with the IGF-I receptor. This structural difference between insulin and IGF-I gives each a higher binding affinity for its own receptor.
Geddes
S
S
School of Crystallography, Birkbeck College, University of London, Malet Street, London WC1 7HX, UK.
Holst
P
P
Grotzinger
J
J
Gill
R
R
Nugent
P
P
De Meyts
P
P
Wollmer
A
A
Wood
S
S
Pitts
J
J
D016475:3T3 Cells; D017667:Adipocytes / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002942:Circular Dichroism; D007334:Insulin-Like Growth Factor I / Q000031:analogs & derivatives* / Q000737:chemistry; D007335:Insulin-Like Growth Factor II / Q000737:chemistry; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D051381:Rats; D017526:Receptor, IGF Type 1 / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D013329:Structure-Activity Relationship
D002352:Carrier Proteins; D010455:Peptides; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II; D017526:Receptor, IGF Type 1