Monitoring the prevention of amyloid fibril formation by α‐crystallin

Abstract

The molecular chaperone, alpha-crystallin, has the ability to prevent the fibrillar aggregation of proteins implicated in human diseases, for example, amyloid beta peptide and alpha-synuclein. In this study, we examine, in detail, two aspects of alpha-crystallin's fibril-suppressing ability: (a) its temperature dependence, and (b) the nature of the aggregating species with which it interacts. First, the efficiency of alpha-crystallin to suppress fibril formation in kappa-casein and alpha-synuclein increases with temperature, despite their rate of fibrillation also increasing in the absence of alpha-crystallin. This is consistent with an increased chaperone ability of alpha-crystallin at higher temperatures to protect target proteins from amorphous aggregation [GB Reddy, KP Das, JM Petrash & WK Surewicz (2000) J Biol Chem275, 4565-4570]. Second, dual polarization interferometry was used to monitor real-time alpha-synuclein aggregation in the presence and absence of alphaB-crystallin. In contrast to more common methods for monitoring the time-dependent formation of amyloid fibrils (e.g. the binding of dyes like thioflavin T), dual polarization interferometry data did not reveal any initial lag phase, generally attributed to the formation of prefibrillar aggregates. It was shown that alphaB-crystallin interrupted alpha-synuclein aggregation at its earliest stages, most likely by binding to partially folded monomers and thereby preventing their aggregation into fibrillar structures.

The functional roles of the unstructured N- and C-terminal regions in αB-crystallin and other mammalian small heat-shock proteins

“…The interaction of αBc with amyloid fibril-forming proteins, e.g. α-synuclein, ataxin-3, apolipoprotein C-II, kappa-casein and β2-microgobulin, is such a situation (Cox et al 2016;Esposito et al 2013;Hatters et al 2001;Rekas et al 2004;Rekas et al 2007;Robertson et al 2010). The variation in sHsp chaperone mechanism depending on conditions and the degree of unfolding of the target protein is consistent with various studies.…”

Small heat-shock proteins: important players in regulating cellular proteostasis

“…Thus, the design of such studies does not address the effect(s) sHsps have on the latter stages of aggregation, which is as important to consider since, in vivo, levels of sHsps in the cell increase after aggregation has commenced as a result of the activation of the stress response [104,105]. Apart from interacting with monomeric species, sHsps also bind to species formed further along the aggregation pathway, including mature amyloid fibrils [106][107][108][109]. For example, when introduced during the elongation phase of α-synuclein (α-syn) or amyloid- peptide (A aggregation, αBc prevents further fibril growth by binding along the length of mature fibrils, preventing secondary nucleation events that facilitate further fibril growth [106][107][108].…”

“…Apart from interacting with monomeric species, sHsps also bind to species formed further along the aggregation pathway, including mature amyloid fibrils [106][107][108][109]. For example, when introduced during the elongation phase of α-synuclein (α-syn) or amyloid- peptide (A aggregation, αBc prevents further fibril growth by binding along the length of mature fibrils, preventing secondary nucleation events that facilitate further fibril growth [106][107][108]. Our recent work, using apolipoprotein C-II (apoC-II) as a model fibril-forming protein, has shown that, by binding to fibrils, αBc stabilises them, preventing their (dilution-induced) fragmentation, and causes them to associate (tangle) into larger species reminiscent of protein inclusions [109].…”

“…Both PD and Huntington's disease are intracellular protein aggregation diseases, like CJD. We have previously demonstrated that Bc prevents the aggregation of -syn, the principal protein in Lewy body deposits, and that Bc also binds to intact -syn fibrils 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 to prevent their further growth [106,107,167]. Furthermore, several sHsps (Hsp20, Hsp22, HspB7 and HspB9) inhibit the aggregation of the polyQ huntingtin protein responsible for Huntington disease and also protect against cell death triggered by the deposition of the aberrant protein [173,196].…”

PAMAM Dendrimers as Potential Agents against Fibrillation ofα‐Synuclein, a Parkinson's Disease‐Related Protein

“…Decreased TFT binding could also potentially result from agglomeration of a-synuclein fibrils, [22] , but such an effect was previously seen in the final stages of the fibrillation process and was preceded by a rapid growth in TFT fluorescence, which was not observed in the presence of PAMAM.…”