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

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dc.contributor.authorForbes, B.
dc.contributor.authorTurner, D.
dc.contributor.authorHodge, S.
dc.contributor.authorMcNeil, K.
dc.contributor.authorForsberg, G.
dc.contributor.authorWallace, J.
dc.date.issued1998
dc.description.abstractWe 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.
dc.identifier.citationJournal of Biological Chemistry, 1998; 273(8):4647-4652
dc.identifier.doi10.1074/jbc.273.8.4647
dc.identifier.issn0021-9258
dc.identifier.issn1083-351X
dc.identifier.orcidHodge, S. [0000-0002-3602-9927] [0000-0002-9401-298X]
dc.identifier.urihttp://hdl.handle.net/2440/11309
dc.language.isoen
dc.publisherAMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
dc.source.urihttps://doi.org/10.1074/jbc.273.8.4647
dc.subjectAnimals
dc.subjectCattle
dc.subjectDisulfides
dc.subjectTrypsin
dc.subjectSomatomedins
dc.subjectInsulin-Like Growth Factor Binding Protein 2
dc.subjectChromatography, High Pressure Liquid
dc.subjectPeptide Mapping
dc.subjectMutagenesis
dc.subjectSequence Deletion
dc.subjectBinding Sites
dc.subjectAmino Acid Sequence
dc.subjectMolecular Sequence Data
dc.titleLocalization 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
dc.typeJournal article
pubs.publication-statusPublished

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