The formation of a non-covalent complex between mature transforming growth factor β1 (TGF-β1) and its pro region, the β1-latency-associated peptide (β1-LAP), is important in regulating the activity of this multipotent growth factor. We have overexpressed simian β1-LAP in Chinese hamster ovary (CHO) cells to produce a cell line which secretes β1-LAP into the culture medium at > 1 mg/l, thus enabling structural studies of complex formation between β1-LAP and TGF-β1. The simian β1-LAP expressed in CHO cells reversed the growth inhibitory effect of exogenous TGF-β1 on Mv1Lu (mink lung epithelial) cells and was able to form a cross-linked complex with 125I-TGF-β1. Simian β1-LAP was purified to homogeneity by a combination of ammonium sulphate precipitation, gel filtration, dye ligand chromatography and anion-exchange chromatography, with a yield of 15%. The purified protein had an apparent molecular mass of 114 kDa as determined by SDS/PAGE, which is greater than that determined for the transient expression of simian β1-LAP in COS-1 and for the simian precursor of TGF-β1 (pro-TGF-β1) in CHO cells, this major difference being due to more extensive glycosylation of β1-LAP expressed by this CHO clone. Far-UV CD spectroscopy of simian β1-LAP indicates a mostly β-sheet structure, with extensive structural rearrangements occurring upon formation of the latent complex between TGF-β1 and β1-LAP.
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January 1996
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Research Article|
January 01 1996
Structural characterization of the latent complex between transforming growth factor β1 and β1-latency-associated peptide Available to Purchase
John D. DIGNAM;
John D. DIGNAM
1Biochemistry and Molecular Biology, Medical College of Ohio, 3000 Arlington Avenue, Toledo, OH 43699-0008, U.S.A.
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Larry E. GENTRY
Larry E. GENTRY
†
†To whom correspondence should be addressed.
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Publisher: Portland Press Ltd
Received:
June 12 1995
Revision Received:
August 14 1995
Accepted:
August 17 1995
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London © 1996
1996
Biochem J (1996) 313 (1): 343–351.
Article history
Received:
June 12 1995
Revision Received:
August 14 1995
Accepted:
August 17 1995
Citation
Grainne A. McMAHON, John D. DIGNAM, Larry E. GENTRY; Structural characterization of the latent complex between transforming growth factor β1 and β1-latency-associated peptide. Biochem J 1 January 1996; 313 (1): 343–351. doi: https://doi.org/10.1042/bj3130343
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