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https://hdl.handle.net/2440/111715
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Type: | Journal article |
Title: | The influence of the N-terminal region proximal to the core domain on the assembly and chaperone activity of αB-crystallin |
Author: | Jovcevski, B. Andrew Aquilina, J. Benesch, J. Ecroyd, H. |
Citation: | Cell Stress and Chaperones, 2018; 23(5):827-836 |
Publisher: | Springer |
Issue Date: | 2018 |
ISSN: | 1355-8145 1466-1268 |
Statement of Responsibility: | Blagojce Jovcevski, J. Andrew Aquilina, Justin L.P. Benesch, Heath Ecroyd |
Abstract: | αB-Crystallin (HSPB5) is a small heat-shock protein that is composed of dimers that then assemble into a polydisperse ensemble of oligomers. Oligomerisation is mediated by heterologous interactions between the C-terminal tail of one dimer and the core "α-crystallin" domain of another and stabilised by interactions made by the N-terminal region. Comparatively little is known about the latter contribution, but previous studies have suggested that residues in the region 54-60 form contacts that stabilise the assembly. We have generated mutations in this region (P58A, S59A, S59K, R56S/S59R and an inversion of residues 54-60) to examine their impact on oligomerisation and chaperone activity in vitro. By using native mass spectrometry, we found that all the αB-crystallin mutants were assembly competent, populating similar oligomeric distributions to wild-type, ranging from 16-mers to 30-mers. However, circular dichroism spectroscopy, intrinsic tryptophan and bis-ANS fluorescence studies demonstrated that the secondary structure differs to wild type, the 54-60 inversion mutation having the greatest impact. All the mutants exhibited a dramatic decrease in exposed hydrophobicity. We also found that the mutants in general were equally active as the wild-type protein in inhibiting the amorphous aggregation of insulin and seeded amyloid fibrillation of α-synuclein in vitro, except for the 54-60 inversion mutant, which was significantly less effective at inhibiting insulin aggregation. Our data indicate that alterations in the part of the N-terminal region proximal to the core domain do not drastically affect the oligomerisation of αB-crystallin, reinforcing the robustness of αB-crystallin in functioning as a molecular chaperone. |
Keywords: | Proteostasis; molecular chaperone; small heat-shock protein; native mass spectrometry; protein aggregation; αB-crystallin; HSPB5; amyloid fibrils |
Rights: | © Cell Stress Society International 2018 |
DOI: | 10.1007/s12192-018-0889-y |
Grant ID: | http://purl.org/au-research/grants/arc/FT110100586 http://purl.org/au-research/grants/arc/LE0882289 |
Published version: | http://dx.doi.org/10.1007/s12192-018-0889-y |
Appears in Collections: | Aurora harvest 8 Biochemistry publications |
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