Localization of an insulin-like growth factor (IGF) binding site of bovine IGF binding protein-2 using disulfide mapping and deletion mutation analysis of the C-terminal domain.
J Biol Chem
; 273(8): 4647-52, 1998 Feb 20.
Article
en En
| MEDLINE
| ID: mdl-9468524
ABSTRACT
We have investigated which region(s) of bovine insulin-like growth factor binding protein-2 (bIGFBP-2) interact with insulin-like growth factors (IGFs) using C-terminally truncated forms of bIGFBP-2. Initially to aid in mutant design, we defined the disulfide bonding pattern of bIGFBP-2 C-terminal region using enzymatic digestion. The pattern is Cys186-Cys220, Cys231-Cys242, and Cys244-Cys265. In addition, cyanogen bromide cleavage of bIGFBP-2 revealed that the N- and C-terminal cysteine-rich domains were not linked by disulfide bonds. Taking the disulfide bonding pattern into consideration, C-terminal truncation mutants were designed and expressed in COS-1 mammalian cells. Following IGF binding assays, a region between residues 222 and 236 was identified as important in IGF binding. Specifically, mutants truncated by 14, 36, and 48 residues from the C terminus bound IGFs to the same extent as wild type (WT) bIGFBP-2. Removal of 63 residues resulted in a greatly reduced (up to 80-fold) ability to bind IGF compared with WT bIGFBP-2. Interestingly this mutant lacked the IGF-II binding preference of WT bIGFBP-2. Residues 236-270 also appeared to play a role in determining IGF binding specificity as their removal resulted in mutants with higher IGF-II binding affinity.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Somatomedinas
/
Proteína 2 de Unión a Factor de Crecimiento Similar a la Insulina
/
Disulfuros
Tipo de estudio:
Prognostic_studies
Límite:
Animals
Idioma:
En
Revista:
J Biol Chem
Año:
1998
Tipo del documento:
Article
País de afiliación:
Australia