Abstract
Nine neurodegenerative disorders, called polyglutamine (polyQ) diseases, are characterized by the formation of intranuclear amyloid-like aggregates by nine proteins containing a polyQ tract above a threshold length. These insoluble aggregates and/or some of their soluble precursors are thought to play a role in the pathogenesis. The mechanism by which polyQ expansions trigger the aggregation of the relevant proteins remains, however, unclear. In this work, polyQ tracts of different lengths were inserted into a solvent-exposed loop of the β-lactamase BlaP and the effects of these insertions on the properties of BlaP were investigated by a range of biophysical techniques. The insertion of up to 79 glutamines does not modify the structure of BlaP; it does, however, significantly destabilize the enzyme. The extent of destabilization is largely independent of the polyQ length, allowing us to study independently the effects intrinsic to the polyQ length and those related to the structural integrity of BlaP on the aggregating properties of the chimeras. Only chimeras with 55Q and 79Q readily form amyloid-like fibrils; therefore, similarly to the proteins associated with diseases, there is a threshold number of glutamines above which the chimeras aggregate into amyloid-like fibrils. Most importantly, the chimera containing 79Q forms amyloid-like fibrils at the same rate whether BlaP is folded or not, whereas the 55Q chimera aggregates into amyloid-like fibrils only if BlaP is unfolded. The threshold value for amyloid-like fibril formation depends, therefore, on the structural integrity of the β-lactamase moiety and thus on the steric and/or conformational constraints applied to the polyQ tract. These constraints have, however, no significant effect on the propensity of the 79Q tract to trigger fibril formation. These results suggest that the influence of the protein context on the aggregating properties of polyQ disease-associated proteins could be negligible when the latter contain particularly long polyQ tracts.
Highlights
Polyglutamine diseases are neurodegenerative disorders caused by the expansion of unstable CAG trinucleotide repeats in the translated region of unrelated genes
This work clearly demonstrates that the threshold number of glutamines above which the chimeric proteins aggregate into amyloid-like fibrils critically depends on the structural integrity of the b-lactamase moiety and on the constraints applied to the polyQ tract
It suggests that the modulating effects of the protein context on the aggregating properties of proteins associated with polyQ diseases could be negligible when the latter contain long polyQ tracts
Summary
Polyglutamine (polyQ) diseases are neurodegenerative disorders caused by the expansion of unstable CAG trinucleotide repeats in the translated region of unrelated genes. These CAG repeats encode a polyglutamine stretch in the corresponding proteins [1]. There is a threshold in the number of repeats above which polyQ proteins become pathogenic The value of this threshold varies from one disease to another but generally resides between 35 and 45 glutamines [5]. The mechanism of toxicity associated with pathological expansion of polyglutamine tracts remains unclear, a large body of evidence indicates that it is associated with protein misfolding and aggregation [7,8]. The cytotoxicity of proteins containing an expanded polyQ tract has been attributed to: (i) the formation of inclusion bodies [9,10], (ii) the presence of misfolded protein monomers [11] and (iii) the transient formation of oligomers during the aggregation process [12,13,14]
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