Prevention of amyloid fibril formation of amyloidogenic chicken cystatin by site-specific glycosylation in yeast.

Asn106-Thr108) was introduced by site-directed mutagenesis into the wild-type and amyloidogenic chicken cystatins to construct the glycosylated form of chicken cystatins. Both the glycosylated and unglycosylated forms of wild-type and amyloidogenic mutant I66Q cystatin were expressed and secreted in a culture medium of yeast Pichia pastoris transformants. Comparison of the amount of insoluble aggregate, the secondary structure, and fibrillogenicity has shown that the N-linked glycosylation could prevent amyloid fibril formation of amyloidogenic chicken cystatin secreted in yeast cells without affecting its inhibitory activities. Further study showed this glycosylation could inhibit the formation of cystatin dimers. Therefore, our data strongly suggested that the mechanism causing the prevention of amyloidogenic cystation fibril formation may be realized through suppression of the formation of three-dimensional domain-swapped dimers and oligomers of amyloidogenic cystatin by the glycosylated chains at position 106. -->
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Substance Nomenclature: 0 (Amyloid)
0 (Cystatins)
0 (Cysteine Proteinase Inhibitors)
0 (Recombinant Proteins)
0 (cystatin, egg-white)
Entry Date(s): Date Created: 20060126 Date Completed: 20060314 Latest Revision: 20181113
Update Code: 20231215
PubMed Central ID: PMC2242452
DOI: 10.1110/ps.051753306
PMID: 16434741
Autor: He J; Department of Biological Chemistry, Yamaguchi University, Yamaguchi 753-8515, Japan., Song Y, Ueyama N, Saito A, Azakami H, Kato A
Jazyk: angličtina
Zdroj: Protein science : a publication of the Protein Society [Protein Sci] 2006 Feb; Vol. 15 (2), pp. 213-22.
DOI: 10.1110/ps.051753306
Abstrakt: To address the role of glycosylation on fibrillogenicity of amyloidogenic chicken cystatin, the consensus sequence for N-linked glycosylation (Asn106-Ile108 --> Asn106-Thr108) was introduced by site-directed mutagenesis into the wild-type and amyloidogenic chicken cystatins to construct the glycosylated form of chicken cystatins. Both the glycosylated and unglycosylated forms of wild-type and amyloidogenic mutant I66Q cystatin were expressed and secreted in a culture medium of yeast Pichia pastoris transformants. Comparison of the amount of insoluble aggregate, the secondary structure, and fibrillogenicity has shown that the N-linked glycosylation could prevent amyloid fibril formation of amyloidogenic chicken cystatin secreted in yeast cells without affecting its inhibitory activities. Further study showed this glycosylation could inhibit the formation of cystatin dimers. Therefore, our data strongly suggested that the mechanism causing the prevention of amyloidogenic cystation fibril formation may be realized through suppression of the formation of three-dimensional domain-swapped dimers and oligomers of amyloidogenic cystatin by the glycosylated chains at position 106.
Databáze: MEDLINE
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