Human bone morphogenetic protein-2 (hBMP-2) characterization by physical-chemical, immunological and biological assays.
BMP-2
C2C12 bioassay
CHO cell-derived
Efficient osteoinductor
Escherichia coli-derived
Journal
AMB Express
ISSN: 2191-0855
Titre abrégé: AMB Express
Pays: Germany
ID NLM: 101561785
Informations de publication
Date de publication:
17 Feb 2020
17 Feb 2020
Historique:
received:
07
11
2019
accepted:
27
01
2020
entrez:
19
2
2020
pubmed:
19
2
2020
medline:
19
2
2020
Statut:
epublish
Résumé
Commercially available preparations of methionyl-human BMP-2 and CHO-derived hBMP-2, which belongs to the transforming growth factor β (TGF-β) superfamily, were used for a complete characterization. This protein is an extremely efficient osteoinductor that plays an important role during bone regeneration and embryonic development. Characterization was carried out via SDS-PAGE and Western blotting, followed by reversed-phase HPLC, size-exclusion HPLC and MALDI-TOF-MS. The classical in vitro bioassay, based on the induction of alkaline phosphatase activity in C2C12 cells, confirmed that hBMP-2 biological activity is mostly related to the dimeric form, being ~ 4-fold higher for the CHO-derived glycosylated form when compared with the E. coli counterpart. The E. coli-derived met-hBMP-2 has shown, by MALDI-TOF-MS, a large presence of the bioactive dimer. A more complex molecular mass (MM) distribution was found for the CHO-derived product, whose exact MM has never been reported because of its variable glycosylation. A method based on RP-HPLC was set up, allowing a quantitative and qualitative hBMP-2 determination even directly on ongoing culture media. Considering that hBMP-2 is highly unstable, presenting moreover an extremely high aggregate value, we believe that these data pave the way to a necessary characterization of this important factor when synthesized by DNA recombinant techniques in different types of hosts.
Identifiants
pubmed: 32067115
doi: 10.1186/s13568-020-0964-5
pii: 10.1186/s13568-020-0964-5
pmc: PMC7026339
doi:
Types de publication
Journal Article
Langues
eng
Pagination
34Subventions
Organisme : Fundação de Amparo à Pesquisa do Estado de São Paulo
ID : 2015/15446-0
Organisme : Fundação de Amparo à Pesquisa do Estado de São Paulo
ID : 2016/24724-6
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