Amyloid fibril formation from full-length and fragments of amylin.
Amyloiddeposits of fibrillar human amylin (hA) in the pancreas may be a causative factor in type-2 diabetes. A detailed comparison of in vitro fibril formation by full-length hA(1-37) versus fragments of this peptide-hA(8-37) and hA(20-29)-is presented. Circular dichroism spectroscopy revealed that fibril formation was accompanied by a conformational change: random coil to beta-sheet/alpha-helical structure. Fibril morphologies were visualized by electron microscopy and displayed a remarkable diversity. hA(20-29) formed flat ribbons consisting of numerous 3. 6-nm-wide protofibrils. In contrast, hA(1-37) and hA(8-37) formed polymorphic higher order fibrils by lateral association and/or coiling together of 5.0-nm-wide protofibril subunits. For full-length hA(1-37), the predominant fibril type contained three protofibrils and for hA(8-37), the predominant type contained two protofibrils. Polymerization was also monitored with the thioflavin-T binding assay, which revealed different kinetics of assembly for hA(1-37) and hA(8-37) fibrils. hA(20-29) fibrils did not bind thioflavin-T. Together the results demonstrate that the N-terminal region of the hA peptide influences the relative frequencies of the various higher order fibril types and thereby the overall kinetics of fibril formation. Furthermore, while residues 20-29 contribute to the fibrils' beta-sheet core, the flanking C- and N-terminal regions of the hA peptide determine the interactions involved in the formation of higher order coiled polymorphic superstructures.
Department of Biophysical Chemistry, M. E. Müller Institute for Structural Biology, Basel, Switzerland.
C
C
Goldsbury
K
K
Goldie
J
J
Pellaud
J
J
Seelig
P
P
Frey
S A
SA
Müller
J
J
Kistler
G J
GJ
Cooper
U
U
Aebi
D000682:Amyloid / Q000378:metabolism* / Q000648:ultrastructure*; D000818:Animals; D001483:Base Sequence; D052160:Benzothiazoles; D002942:Circular Dichroism; D005456:Fluorescent Dyes; D006801:Humans; D058228:Islet Amyloid Polypeptide; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments; D017433:Protein Structure, Secondary; D051381:Rats; D016415:Sequence Alignment; D013844:Thiazoles
D000682:Amyloid; D052160:Benzothiazoles; D005456:Fluorescent Dyes; D058228:Islet Amyloid Polypeptide; D010446:Peptide Fragments; D013844:Thiazoles; C009462:thioflavin T