A novel assay in vitro of human islet amyloid polypeptide amyloidogenesis and effects of insulin secretory vesicle peptides on amyloid formation

Yogish C Kudva, Cheryl Mueske, Peter C. Butler, Norman L. Eberhardt

Research output: Contribution to journalArticle

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Abstract

Human islet amyloid polypeptide (IAPP) is a 37-residue peptide that is co-secreted with insulin by the β-cell and might be involved in the pathogenesis of non-insulin-dependent diabetes mellitus. We developed an improved assay in vitro based on the fluorescence of bound thioflavin T to study factors affecting amyloidogenesis. Monomeric IAPP formed amyloid fibrils, as detected by increased fluorescence and by electron microscopy. Fluorimetric analysis revealed that the initial rate of amyloid formation was: (1) proportional to the peptide monomer concentration, (2) maximal at pH 9.5, (3) maximal at 200 mM KCl, and (4) proportional to temperature from 4 to 37°C. We found that 5-fold and 10-fold molar excesses of proinsulin inhibited fibril formation by 39% and 59% respectively. Insulin was somewhat more potent with 5-fold and 10-fold molar excesses inhibiting fibril formation by 69% and 73% respectively, whereas C-peptide had no effect at these concentrations. Thus at physiological ratios of IAPP to insulin, insulin and proinsulin, but not C-peptide, can retard amyloidogenesis. Because insulin resistance or hyperglycaemia increase the IAPP-to-insulin ratio, increased intracellular IAPP compared with insulin expression in genetically predisposed individuals might contribute to intracellular amyloid formation, β-cell death and the genesis of non-insulin-dependent diabetes mellitus.

Original languageEnglish (US)
Pages (from-to)809-813
Number of pages5
JournalBiochemical Journal
Volume331
Issue number3
StatePublished - May 1 1998

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Islet Amyloid Polypeptide
Secretory Vesicles
Amyloid
Assays
Insulin
Peptides
C-Peptide
Proinsulin
Type 2 Diabetes Mellitus
Medical problems
Fluorescence
Fluorescence Microscopy
Hyperglycemia
Cell death
Insulin Resistance
In Vitro Techniques
Electron Microscopy
Cell Death
Electron microscopy
Monomers

ASJC Scopus subject areas

  • Biochemistry

Cite this

A novel assay in vitro of human islet amyloid polypeptide amyloidogenesis and effects of insulin secretory vesicle peptides on amyloid formation. / Kudva, Yogish C; Mueske, Cheryl; Butler, Peter C.; Eberhardt, Norman L.

In: Biochemical Journal, Vol. 331, No. 3, 01.05.1998, p. 809-813.

Research output: Contribution to journalArticle

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