Recombinant platelet-derived growth factor (PDGF)-BB was expressed and secreted from yeast in order to study the structure-function relationships of this mitogen. A simple purification scheme has been developed which yields greater than 95% pure PDGF-BB. Analysis of this recombinant PDGF-BB shows partial proteolysis after arginine-32. Substitution of this arginine residue, or arginine-28 [a potential KEX2 (lysine-arginine endopeptidase) cleavage site], prevents or reduces cleavage of PDGF-BB respectively. These mutations result in a 5-fold increase in expression levels of PDGF-BB, and the resulting mutant proteins show higher activity in a number of biological assays than the cleaved wildtype PDGF-BB. These data are in accord with previous work by Giese, LaRochelle, May-Siroff, Robbins & Aaronson [(1990) Mol. Cell Biol. 10, 5496-5501] suggesting that the region isoleucine-25-phenylalanine-37 is involved in PDGF-receptor binding.
Purification and analysis of proteinase-resistant mutants of recombinant platelet-derived growth factor-BB exhibiting improved biological activity
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A L Cook, P M Kirwin, S Craig, L J Bawden, D R Green, M J Price, S J Richardson, A Fallon, A H Drummond, R M Edwards, J M Clements; Purification and analysis of proteinase-resistant mutants of recombinant platelet-derived growth factor-BB exhibiting improved biological activity. Biochem J 1 January 1992; 281 (1): 57–65. doi: https://doi.org/10.1042/bj2810057
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