ISSN:
0006-3592
Keywords:
furin
;
proprotein convertases
;
transforming growth factor beta
;
baculovirus
;
precursor proteins
;
overexpression systems
;
growth factors
;
Chemistry
;
Biochemistry and Biotechnology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Process Engineering, Biotechnology, Nutrition Technology
Notes:
One important limitation of the widely used insect baculovirus overexpression system is its inefficiency to properly process heterologous proteins which are initially biosynthesized as larger inactive precursor proteins. One example is transforming growth factor beta 1 (TGFβ1), a 25-kDa homodimeric protein with pleiotropic functions. As many growth factors, the inactive TGFβ1 precursor molecule needs to be proteolytically cleaved C-terminal to a basic sequence to yield the mature and active homodimer. In insect cells, a large proportion of overexpressed TGFβ1 was found in an inactive precursor form suggesting that the levels of endogenous convertases are limiting for the production of mature and bioactive TGFβ1 in this system. We have demonstrated that furin, a member of a novel family of mammalian prohormone convertases (PCs) can efficiently process TGFβ1 precursor resulting in the production of the mature and active growth factor. Taking advantage of this observation, we have developed an improved overproduction system for TGFβ1 by coexpressing prohTGFβ1 and human furin convertase in High Five cells. Using this system, the production of mature active TGFβ1 increased in a dose-dependent fashion reaching up to 7.8-fold the amount obtained with the growth factor only. Thus, eliminating the rate-limiting step in recombinant TGFβ1 production maximizes its processing efficiency and the yield of the mature active growth factor. Such simple and efficient technology could be useful for large scale production of other proproteins which undergo similar maturation processes and share furin recognition sequences at the junction between the proregion and the mature polypeptide. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 58:85-91, 1998.
Additional Material:
5 Ill.
Type of Medium:
Electronic Resource
