The C-terminal domain of the antiamyloid chaperone DNAJB6 binds to amyloid-β peptide fibrils and inhibits secondary nucleation
Österlund, Nicklas ; Frankel, Rebecca LU ; Carlsson, Andreas LU ; Thacker, Dev LU ; Karlsson, Maja LU ; Matus, Vanessa ; Gräslund, Astrid ; Emanuelsson, Cecilia LU
and Linse, Sara
LU
(2023)
In Journal of Biological Chemistry
299(11).
- Abstract
The DNAJB6 chaperone inhibits fibril formation of aggregation-prone client peptides through interaction with aggregated and oligomeric forms of the amyloid peptides. Here, we studied the role of its C-terminal domain (CTD) using constructs comprising either the entire CTD or the first two or all four of the CTD β-strands grafted onto a scaffold protein. Each construct was expressed as WT and as a variant with alanines replacing five highly conserved and functionally important serine and threonine residues in the first β-strand. We investigated the stability, oligomerization, antiamyloid activity, and affinity for amyloid-β (Aβ42) species using optical spectroscopy, native mass spectrometry, chemical crosslinking, and surface plasmon... (More)
The DNAJB6 chaperone inhibits fibril formation of aggregation-prone client peptides through interaction with aggregated and oligomeric forms of the amyloid peptides. Here, we studied the role of its C-terminal domain (CTD) using constructs comprising either the entire CTD or the first two or all four of the CTD β-strands grafted onto a scaffold protein. Each construct was expressed as WT and as a variant with alanines replacing five highly conserved and functionally important serine and threonine residues in the first β-strand. We investigated the stability, oligomerization, antiamyloid activity, and affinity for amyloid-β (Aβ42) species using optical spectroscopy, native mass spectrometry, chemical crosslinking, and surface plasmon resonance technology. While DNAJB6 forms large and polydisperse oligomers, CTD was found to form only monomers, dimers, and tetramers of low affinity. Kinetic analyses showed a shift in inhibition mechanism. Whereas full-length DNAJB6 activity is dependent on the serine and threonine residues and efficiently inhibits primary and secondary nucleation, all CTD constructs inhibit secondary nucleation only, independently of the serine and threonine residues, although their dimerization and thermal stabilities are reduced by alanine substitution. While the full-length DNAJB6 inhibition of primary nucleation is related to its propensity to form coaggregates with Aβ, the CTD constructs instead bind to Aβ42 fibrils, which affects the nucleation events at the fibril surface. The retardation of secondary nucleation by DNAJB6 can thus be ascribed to the first two β-strands of its CTD, whereas the inhibition of primary nucleation is dependent on the entire protein or regions outside the CTD.
(Less)
- author
- Österlund, Nicklas
; Frankel, Rebecca
LU
; Carlsson, Andreas
LU
; Thacker, Dev
LU
; Karlsson, Maja
LU
; Matus, Vanessa
; Gräslund, Astrid
; Emanuelsson, Cecilia
LU
and Linse, Sara
LU
- organization
- publishing date
- 2023
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- amyloid-beta, chaperone DNAJ, neurodegeneration, protein aggregation, protein engineering, self-assembly
- in
- Journal of Biological Chemistry
- volume
- 299
- issue
- 11
- article number
- 105317
- publisher
- American Society for Biochemistry and Molecular Biology
- external identifiers
-
- pmid:37797698
- scopus:85175246202
- ISSN
- 0021-9258
- DOI
- 10.1016/j.jbc.2023.105317
- language
- English
- LU publication?
- yes
- id
- 7b1f6f09-0960-476a-8f8b-01c34dc3fbdb
- date added to LUP
- 2023-12-11 12:28:58
- date last changed
- 2024-04-24 05:58:36
@article{7b1f6f09-0960-476a-8f8b-01c34dc3fbdb,
abstract = {{<p>The DNAJB6 chaperone inhibits fibril formation of aggregation-prone client peptides through interaction with aggregated and oligomeric forms of the amyloid peptides. Here, we studied the role of its C-terminal domain (CTD) using constructs comprising either the entire CTD or the first two or all four of the CTD β-strands grafted onto a scaffold protein. Each construct was expressed as WT and as a variant with alanines replacing five highly conserved and functionally important serine and threonine residues in the first β-strand. We investigated the stability, oligomerization, antiamyloid activity, and affinity for amyloid-β (Aβ42) species using optical spectroscopy, native mass spectrometry, chemical crosslinking, and surface plasmon resonance technology. While DNAJB6 forms large and polydisperse oligomers, CTD was found to form only monomers, dimers, and tetramers of low affinity. Kinetic analyses showed a shift in inhibition mechanism. Whereas full-length DNAJB6 activity is dependent on the serine and threonine residues and efficiently inhibits primary and secondary nucleation, all CTD constructs inhibit secondary nucleation only, independently of the serine and threonine residues, although their dimerization and thermal stabilities are reduced by alanine substitution. While the full-length DNAJB6 inhibition of primary nucleation is related to its propensity to form coaggregates with Aβ, the CTD constructs instead bind to Aβ42 fibrils, which affects the nucleation events at the fibril surface. The retardation of secondary nucleation by DNAJB6 can thus be ascribed to the first two β-strands of its CTD, whereas the inhibition of primary nucleation is dependent on the entire protein or regions outside the CTD.</p>}},
author = {{Österlund, Nicklas and Frankel, Rebecca and Carlsson, Andreas and Thacker, Dev and Karlsson, Maja and Matus, Vanessa and Gräslund, Astrid and Emanuelsson, Cecilia and Linse, Sara}},
issn = {{0021-9258}},
keywords = {{amyloid-beta; chaperone DNAJ; neurodegeneration; protein aggregation; protein engineering; self-assembly}},
language = {{eng}},
number = {{11}},
publisher = {{American Society for Biochemistry and Molecular Biology}},
series = {{Journal of Biological Chemistry}},
title = {{The C-terminal domain of the antiamyloid chaperone DNAJB6 binds to amyloid-β peptide fibrils and inhibits secondary nucleation}},
url = {{http://dx.doi.org/10.1016/j.jbc.2023.105317}},
doi = {{10.1016/j.jbc.2023.105317}},
volume = {{299}},
year = {{2023}},
}