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Analysis of core region from egg white lysozyme forming amyloid fibrils.

Tokunaga Y, Sakakibara Y, Kamada Y, Watanabe K, Sugimoto Y - Int. J. Biol. Sci. (2013)

Bottom Line: The K peptide alone formed definite fibrils having β-sheet structures by incubation of 7 days under acidic conditions at 37°C.A substantial number of fibrils were generated under this pH condition and incubation period.Deletion and substitution of tryptophan in the K peptide resulted in no formation of fibrils.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biochemistry and Bioscience The United Graduate School of Agricultural Sciences, Kagoshima University, Kagoshima 890-0065 Japan.

ABSTRACT
Some of the lysozyme mutants in humans cause systemic amyloidosis. Hen egg white lysozyme (HEWL) has been well studied as a model protein of amyloid fibrils formation. We previously identified an amyloid core region consisting of nine amino acids (designated as the K peptide), which is present at 54-62 in HEWL. The K peptide, with tryptophan at its C- terminus, has the ability of self-aggregation. In the present work we focused on its structural properties in relation to the formation of fibrils. The K peptide alone formed definite fibrils having β-sheet structures by incubation of 7 days under acidic conditions at 37°C. A substantial number of fibrils were generated under this pH condition and incubation period. Deletion and substitution of tryptophan in the K peptide resulted in no formation of fibrils. Tryptophan 62 in lysozyme was suggested to be especially crucial to forming amyloid fibrils. We also show that amyloid fibrils formation of the K peptide requires not only tryptophan 62 but also a certain length containing hydrophobic amino acids. A core region is involved in the significant formation of amyloid fibrils of lysozyme.

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Far-UV CD spectra of the K peptide and other substances. Samples indicated in the legend to Fig. 3 were used. As a control, non-incubated K peptide preparation was also analyzed (K pep-0). Before being subjected to the analysis, each peptide and HEWL was diluted to 80 μg/ml as described in Materials and Methods.
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Figure 4: Far-UV CD spectra of the K peptide and other substances. Samples indicated in the legend to Fig. 3 were used. As a control, non-incubated K peptide preparation was also analyzed (K pep-0). Before being subjected to the analysis, each peptide and HEWL was diluted to 80 μg/ml as described in Materials and Methods.

Mentions: Far UV-CD spectroscopy was conducted to explore the secondary structural features, in particular those of the β-sheet, of the present preparations (Fig. 4). The K peptide incubated at 37°C for 7 days at pH 4 showed a spectrum bearing a minimum at 218 nm (dashed line), which is typical of a β-sheet. The calculation by Yang's method 28 from the spectrum apparently indicated that the content of the β-structure (sum of β-sheet and β-turn) was 87% with a scant random coil, but without an α-helix. As expected, the STDY-K peptide exhibited the signal of the β-sheet (bold dotted line), whose content was calculated to be approximately 70%. Both peptides showed a similar pattern. In contrast, the K peptide before incubation presented an inconstant spectrum having no minimum in the vicinity of 218 nm (long dashed dotted line), indicating that it scarcely had β-conformation. No typical spectrum of CD for the β-structure was given by the peptide GILQINSRG (dotted line), although it was incubated as described above. The solid line in Fig. 4 shows a CD spectrum of HEWL.


Analysis of core region from egg white lysozyme forming amyloid fibrils.

Tokunaga Y, Sakakibara Y, Kamada Y, Watanabe K, Sugimoto Y - Int. J. Biol. Sci. (2013)

Far-UV CD spectra of the K peptide and other substances. Samples indicated in the legend to Fig. 3 were used. As a control, non-incubated K peptide preparation was also analyzed (K pep-0). Before being subjected to the analysis, each peptide and HEWL was diluted to 80 μg/ml as described in Materials and Methods.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3584918&req=5

Figure 4: Far-UV CD spectra of the K peptide and other substances. Samples indicated in the legend to Fig. 3 were used. As a control, non-incubated K peptide preparation was also analyzed (K pep-0). Before being subjected to the analysis, each peptide and HEWL was diluted to 80 μg/ml as described in Materials and Methods.
Mentions: Far UV-CD spectroscopy was conducted to explore the secondary structural features, in particular those of the β-sheet, of the present preparations (Fig. 4). The K peptide incubated at 37°C for 7 days at pH 4 showed a spectrum bearing a minimum at 218 nm (dashed line), which is typical of a β-sheet. The calculation by Yang's method 28 from the spectrum apparently indicated that the content of the β-structure (sum of β-sheet and β-turn) was 87% with a scant random coil, but without an α-helix. As expected, the STDY-K peptide exhibited the signal of the β-sheet (bold dotted line), whose content was calculated to be approximately 70%. Both peptides showed a similar pattern. In contrast, the K peptide before incubation presented an inconstant spectrum having no minimum in the vicinity of 218 nm (long dashed dotted line), indicating that it scarcely had β-conformation. No typical spectrum of CD for the β-structure was given by the peptide GILQINSRG (dotted line), although it was incubated as described above. The solid line in Fig. 4 shows a CD spectrum of HEWL.

Bottom Line: The K peptide alone formed definite fibrils having β-sheet structures by incubation of 7 days under acidic conditions at 37°C.A substantial number of fibrils were generated under this pH condition and incubation period.Deletion and substitution of tryptophan in the K peptide resulted in no formation of fibrils.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biochemistry and Bioscience The United Graduate School of Agricultural Sciences, Kagoshima University, Kagoshima 890-0065 Japan.

ABSTRACT
Some of the lysozyme mutants in humans cause systemic amyloidosis. Hen egg white lysozyme (HEWL) has been well studied as a model protein of amyloid fibrils formation. We previously identified an amyloid core region consisting of nine amino acids (designated as the K peptide), which is present at 54-62 in HEWL. The K peptide, with tryptophan at its C- terminus, has the ability of self-aggregation. In the present work we focused on its structural properties in relation to the formation of fibrils. The K peptide alone formed definite fibrils having β-sheet structures by incubation of 7 days under acidic conditions at 37°C. A substantial number of fibrils were generated under this pH condition and incubation period. Deletion and substitution of tryptophan in the K peptide resulted in no formation of fibrils. Tryptophan 62 in lysozyme was suggested to be especially crucial to forming amyloid fibrils. We also show that amyloid fibrils formation of the K peptide requires not only tryptophan 62 but also a certain length containing hydrophobic amino acids. A core region is involved in the significant formation of amyloid fibrils of lysozyme.

Show MeSH
Related in: MedlinePlus