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Isolation and characterization of ovine IGFBP-4: protein purification and cDNA sequence
journal contribution
posted on 1994-10-01, 00:00 authored by J M Carr, P A Grant, G L Francis, Julie OwensJulie Owens, J C Wallace, P E WaltonThree different molecular mass forms of IGF-binding proteins (IGFBPs) were purified from ovine plasma by IGF-I affinity chromatography and reverse-phase HPLC: a 46 kDa doublet and 29 kDa and 24 kDa forms. Amino-terminal sequence analysis confirmed that these proteins were ovine (o)IGFBP-3 (46 kDa) and two molecular size variants of oIGFBP-4. oIGFBP-3 and the 29 kDa form of oIGFBP-4 were shown to be N-glycosylated. Isoelectric points were determined to be at approximately pH 6 for oIGFBP-3 and at pH 7 and pH 7.5 for the 29 and 24 kDa forms of oIGFBP-4 respectively. The two different molecular mass variants of oIGFBP-4 had similar IGF-binding properties. Compared with human IGFBP-3 and oIGFBP-3, the two variants of oIGFBP-4 exhibited lower relative binding to amino-terminally modified IGF-I analogues in a competitive IGF-binding assay. The full protein sequence of oIGFBP-4, as deduced from the cDNA sequence, showed a high degree of identity with rat (90%), human (96%) and bovine (98%) IGFBP-4. The cDNA sequence also showed homology over regions of the 3' non-coding sequence, particularly in comparison with bovine IGFBP-4 (96%). Northern analysis of mRNA for oIGFBP-4 indicated a 2.6 kb major transcript and two minor transcripts of approximately 2.1 and 1.8 kb. oIGFBP-4 mRNA transcripts were detected in adult ewe liver > kidney > lung > heart and also in several fetal tissues, thus suggesting tissue-specific and developmental regulation. The availability of purified oIGFBP-4 and oIGFBP-3 as well as DNA probes for oIGFBP-4 will enable further study of the properties and functions of these proteins, as well as the establishment of specific assays for these IGFBPs.
History
Journal
Journal of molecular endocrinologyVolume
13Issue
2Pagination
219 - 236Publisher
BioscientificaLocation
Bradley Stok, Eng.Publisher DOI
ISSN
0952-5041Language
engPublication classification
C1.1 Refereed article in a scholarly journalCopyright notice
1994, BioscientificaUsage metrics
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Amino Acid SequenceAnimalsBase SequenceCarrier ProteinsCattleCloning, MolecularDNA, ComplementaryGlycosylationHumansInsulin-Like Growth Factor Binding Protein 4Insulin-Like Growth Factor Binding ProteinsMolecular Sequence DataMolecular WeightRNA, MessengerRatsSequence Homology, Amino AcidSheepSomatomedinsSpecies SpecificityScience & TechnologyLife Sciences & BiomedicineEndocrinology & MetabolismFACTOR-BINDING-PROTEINGROWTH FACTOR-IIGF-IMOLECULAR-CLONINGFACTOR (IGF)-IHUMAN-SERUMRAT SERUMCELL-LINEACIDIDENTIFICATION
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