Aggregation pathway of recombinant human keratinocyte growth factor and its stabilization.
Recombinant human keratinocyte growth factor (rhKGF) is prone to aggregation at elevated temperatures. Its aggregation pathway is proposed to proceed initially with a conformational change which perhaps results from repulsion between positively charged residues in clusters forming heparin binding sites. Unfolding of the protein leads to formation of large soluble aggregates. These soluble aggregates then form disulfide cross-linked precipitates. Finally these precipitates are converted to scrambled disulfides and/or non-disulfide cross-linked precipitates. Stabilizers such as heparin, sulfated polysaccharides, anionic polymers and citrate can greatly decrease the rate of aggregation of rhKGF at elevated temperatures. These molecules may all act by reducing charge repulsion on the protein thus stabilizing the native conformation. EDTA, on the other hand, is found to inhibit disulfide formation in aggregates and has only a moderate stabilizing effect on rhKGF.
Chen
B L
BL
Department of Pharmaceutics and Drug Delivery, Amgen Inc., Thousand Oaks, CA 91320.
Arakawa
T
T
Morris
C F
CF
Kenney
W C
WC
Wells
C M
CM
Pitt
C G
CG
D000838:Anions; D002021:Buffers; D011232:Chemical Precipitation; D002951:Citrates / Q000737:chemistry; D019343:Citric Acid; D004355:Drug Stability; D051526:Fibroblast Growth Factor 10; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors; D006133:Growth Substances / Q000737:chemistry*; D006801:Humans; D011994:Recombinant Proteins / Q000737:chemistry
D000838:Anions; D002021:Buffers; D002951:Citrates; C496453:FGF7 protein, human; D051526:Fibroblast Growth Factor 10; D006133:Growth Substances; D011994:Recombinant Proteins; D051523:Fibroblast Growth Factor 7; D019343:Citric Acid; D005346:Fibroblast Growth Factors