Inhibition of insulin fibrillogenesis with targeted peptides.
Under conditions of acidic pH and elevated temperature, insulin partially unfolds and aggregates into highly structured amyloid fibrils. Aggregation of insulin leads to loss of activity and can trigger an unwanted immune response. Compounds that prevent protein aggregation have been used to stabilize insulin; these compounds generally suppress aggregation only at relatively high inhibitor concentrations. For example, effective inhibition of aggregation of 0.5 mM insulin required arginine concentrations of > or =100 mM. Here, we investigate a targeted approach toward inhibiting insulin aggregation. VEALYL, corresponding to residues B12-17 of full-length insulin, was identified as a short peptide that interacts with full-length insulin. A hybrid peptide was synthesized that contained this binding domain and hexameric arginine; this peptide significantly reduced the rate of insulin aggregation at near-equimolar concentrations. An effective binding domain and N-terminal placement of the arginine hexamer were necessary for inhibitory activity. The data were analyzed using a simple two-step model of aggregation kinetics. These results are useful not only in identifying an insulin aggregation inhibitor but also in extending a targeted protein strategy for modifying aggregation of amyloidogenic proteins.
Gibson
Todd J
TJ
Department of Chemical and Biological Engineering, University of Wisconsin, Madison, Madison, Wisconsin 53706, USA.
Murphy
Regina M
RM
D000595:Amino Acid Sequence; D000682:Amyloid / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000648:ultrastructure; D001665:Binding Sites; D007328:Insulin / Q000378:metabolism*; D007700:Kinetics; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism* / Q000648:ultrastructure; D010455:Peptides; D013329:Structure-Activity Relationship
D000682:Amyloid; D007328:Insulin; D010446:Peptide Fragments; D010455:Peptides