Degradation of islet amyloid polypeptide by neprilysin.
OBJECTIVE
A progressive loss of pancreatic beta cell function, a decrease in beta cell mass and accumulation of islet amyloid is characteristic of type 2 diabetes mellitus. The main constituent of islet amyloid is islet amyloid polypeptide (IAPP). In this study, we examined the ability of the peptidase neprilysin to cleave IAPP and prevent human IAPP-induced pancreatic beta cell toxicity.


METHODS
Neprilysin and a catalytically compromised neprilysin mutant were tested for their ability to inhibit human IAPP fibrillisation and human IAPP-induced pancreatic beta cell cytotoxicity. Degradation of human IAPP by neprilysin was followed by HPLC, and the degradation products were identified by MS.


RESULTS
Neprilysin prevented IAPP fibrillisation by cleaving IAPP at Arg(11)-Leu(12), Leu(12)-Ala(13), Asn(14)-Phe(15), Phe(15)-Leu(16), Asn(22)-Phe(23) and Ala(25)-Ile(26). It also appears to prevent human IAPP fibrillisation through a non-catalytic interaction. Neprilysin protected against beta cell cytotoxicity induced by exogenously added or endogenously produced human IAPP.


CONCLUSIONS
The data presented support a potential therapeutic role for neprilysin in preventing type 2 diabetes mellitus. This study supports the hypothesis that extracellular human IAPP contributes to human IAPP-induced beta cell cytotoxicity. Whether human IAPP exerts its cytotoxic effect through a totally extracellular mechanism or through a cellular reuptake mechanism is unclear at this time.
Department of Molecular and Cellular Biochemistry, University of Kentucky, B236 Biomedical Biological Sciences Research Building, 741 South Limestone Street, Lexington, KY 40536-0509, USA. hguan2@uky.edu
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H
Guan
K M
KM
Chow
R
R
Shah
C J
CJ
Rhodes
L B
LB
Hersh
D000595:Amino Acid Sequence; D000682:Amyloid / Q000378:metabolism; D016564:Amyloid beta-Protein Precursor / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D045744:Cell Line, Tumor; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism* / Q000473:pathology; D004789:Enzyme Activation / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000201:enzymology*; D007340:Insulinoma; D008969:Molecular Sequence Data; D015260:Neprilysin / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D010190:Pancreatic Neoplasms; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D013379:Substrate Specificity / Q000502:physiology; D014162:Transfection
D000682:Amyloid; D016564:Amyloid beta-Protein Precursor; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D015260:Neprilysin