Artigo de peri??dico
Human bone morphogenetic protein???2 (hBMP???2) characterization by physical???chemical, immunological and biological assays
Registro en:
2191-0855
1
10
10.1186/s13568-020-0964-5
0000-0002-6937-1120
0000-0002-7467-3457
53.77
67.00
Autor
SUZUKI, MIRIAM F.
OLIVEIRA, JOAO E.
DAMIANI, RENATA
LIMA, ELIANA R.
AMARAL, KLEICY C.
SANTOS, ANDERSON M. de S.
MAGALH??ES, GERALDO S.
FAVERANI, LEONARDO P.
PEREIRA, LUIS A.V.D.
SILVA, FABIANA M.
BARTOLINI, PAOLO
Resumen
Commercially available preparations of methionyl-human BMP-2 and CHO-derived hBMP-2, which belongs to the
transforming growth factor ?? (TGF-??) 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, sizeexclusion
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 methBMP-
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. Funda????o de Amparo ?? Pesquisa do Estado de S??o Paulo (FAPESP) FAPESP: 15/15446-0; 16/24724-6