Effect of pH and insulin on fibrillogenesis of islet amyloid polypeptide in vitro.
Islet amyloid polypeptide (IAPP) is the constituent of amyloid deposits in pancreatic islets of type 2 diabetes in man, monkeys, and cats. This 37 amino acid peptide aggregates in vitro to form beta-pleated sheet fibrils. Rodent IAPP has a different amino acid sequence and does not form amyloid either in vitro or in vivo. Fibrillogenic properties of human IAPP (hIAPP) were determined in vitro. The effect of pH and time course of fibril formation was studied by light scattering spectroscopy. Aggregation of hIAPP1-37NH2 and hIAPPTyr20-29 (0.25 mg/mL) was maximal at neutral/basic and acidic pH, respectively. The ultrastructure of hIAPP1-37NH2 fibrils (0.2 mg/mL) was examined using negative staining for electron microscopy. Short fibrils composed of 2 or more filaments were observed at pH 3-9 after 30 min incubation. At pH 7-9, IAPP fibrils formed a gel. After 6 months at pH 3, large sheets of parallel fibrils were seen. Specific binding of 125I-hIAPP1-37NH2 to preformed IAPP fibrils detected by quantitative autoradiography and radioassay was maximal at pH 3. Binding was enhanced by insulin (3.7 nmol/L) and unaffected by glucose, calcium, glucagon, and apolipoprotein E. 125I-hIAPP1-37NH2 bound specifically to islet amyloid in pancreatic tissue sections from type 2 diabetic patients.


CONCLUSIONS
Binding to preformed IAPP fibrils is maximal at acid pH when hIAPP is largely in soluble form. IAPP is secreted together with insulin from the acidic secretory granules (pH 5.5) to the neutral pH of the extracellular space under normal conditions. These changes in pH together with increased accumulation of extracellular hIAPP in diabetes may promote amyloid formation.
Chargé
S B
SB
Diabetes Research Laboratories, University of Oxford, Radcliffe Infirmary, U.K.
de Koning
E J
EJ
Clark
A
A
D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure*; D000818:Animals; D002415:Cats; D000882:Haplorhini; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007328:Insulin / Q000494:pharmacology*; D058228:Islet Amyloid Polypeptide; D007515:Islets of Langerhans / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D013053:Spectrophotometry
D000682:Amyloid; D007328:Insulin; D058228:Islet Amyloid Polypeptide