Site-Directed Mutations in the C-Terminal Extension of Human αB-Crystallin Affect Chaperone Function and Block Amyloid Fibril Formation

Treweek, Teresa M.; Ecroyd, Heath; Williams, Danielle M.; Meehan, Sarah; Carver, John A.; Walker, Mark J.

doi:10.1371/journal.pone.0001046

Cited by 42 publications

(54 citation statements)

References 68 publications

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“…Similarly, we found that a double mutation in the C-terminal hinge region of αB-crystallin (i.e. I159A/I161A αB-crystallin) had no effect on its chaperone activity against amorphously aggregating target proteins, but it significantly enhanced its ability to prevent fibril formation by the model proteins κ-casein and ccβ-Trp [106]. Thus, whilst all sHsps share structural features, such as a conserved α-crystallin domain, the differences between the proteins play an important role in their interaction with target proteins.…”

Section: Shsps and The Aβ Peptidesmentioning

confidence: 77%

“…Whether these differences are manifested through distinct target protein binding sites or binding modes remains to be determined. Interestingly, this raises the possibility that mutant forms of sHsps may be designed as more effective inhibitors of fibril formation compared to the wild-type protein and therefore may be an avenue for therapeutic potential in the future [106]. In addition, the variation in chaperone activity of sHsps may provide clues as to the different nature of the partially folded intermediate which are the precursor(s) to disordered (amorphous) versus ordered (amyloid fibril) forms of aggregation.…”

Section: Shsps and The Aβ Peptidesmentioning

confidence: 99%

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Crystallin proteins and amyloid fibrils

Ecroyd

Carver

2008

Cell. Mol. Life Sci.

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220

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Improper protein folding (misfolding) can lead to the formation of disordered (amorphous) or ordered (amyloid fibril) aggregates. The major lens protein, alpha-crystallin, is a member of the small heat-shock protein (sHsp) family of intracellular molecular chaperone proteins that prevent protein aggregation. Whilst the chaperone activity of sHsps against amorphously aggregating proteins has been well studied, its action against fibril-forming proteins has received less attention despite the presence of sHsps in deposits found in fibril-associated diseases (e.g. Alzheimer's and Parkinson's). In this review, the literature on the interaction of alpha B-crystallin and other sHsps with fibril-forming proteins is summarized. In particular, the ability of sHsps to prevent fibril formation, their mechanisms of action and the possible in vivo consequences of such associations are discussed. Finally, the fibril-forming propensity of the crystallin proteins and its implications for cataract formation are described along with the potential use of fibrillar crystallin proteins as bionanomaterials. Improper protein folding (misfolding) can lead to the formation of disordered (amorphous) or ordered (amyloid fibril) aggregates. The major lens protein, α-crystallin, is a member of the small heat-shock protein (sHsp) family of intracellular molecular chaperone proteins that prevent protein aggregation. Whilst the chaperone activity of sHsps against amorphously aggregating proteins has been well studied, its action against fibril-forming proteins has received less attention despite the presence of sHsps in deposits found in fibril-associated diseases (e.g. Alzheimer's and Parkinson's). In this review, the literature on the interaction of αB-crystallin and other sHsps with fibril-forming proteins is summarized. In particular, the ability of sHsps to prevent fibril formation, their mechanisms of action and the possible in vivo consequences of such associations are discussed. Finally, the fibril-forming propensity of the crystallin proteins and its implications for cataract formation are described along with the potential use of fibrillar crystallin proteins as bionanomaterials.

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Section: Shsps and The Aβ Peptidesmentioning

confidence: 77%

Section: Shsps and The Aβ Peptidesmentioning

confidence: 99%

Crystallin proteins and amyloid fibrils

Ecroyd

Carver

2008

Cell. Mol. Life Sci.

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220

234

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“…In vitro, it is well known that C-terminal truncation within the exposed C-terminal extension decreases the solubility of both Ac and Bc [6]. Deamidation occurs extensively to Ac and to Bc with age in the human lens [155] and leads to potential destabilising structural changes due to the introduction of an additional negative charge from the resultant aspartic or glutamic acid sidechains [156].…”

Section: Other Post-translational Modificationsmentioning

confidence: 99%

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

Treweek

Meehan

Ecroyd

et al. 2014

Cell. Mol. Life Sci.

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179

177

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Small heat-shock proteins (sHsps) are a diverse family of intra-cellular molecular chaperone proteins that play a critical role in mitigating and preventing protein aggregation under stress conditions such as elevated temperature, oxidation and infection. In doing so, they assist in the maintenance of protein homeostasis (proteostasis) thereby avoiding the deleterious effects that result from loss of protein function and/or protein aggregation. The chaperone properties of sHsps are therefore employed extensively in many tissues to prevent the development of diseases associated with protein aggregation. Significant progress has been made of late in understanding the structure and chaperone mechanism of sHsps. In this review, we discuss some of these advances, with a focus on mammalian sHsp hetero-oligomerisation, the mechanism by which sHsps act as molecular chaperones to prevent both amorphous and fibrillar protein aggregation, and the role of posttranslational modifications in sHsp chaperone function, particularly in the context of disease. 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 2 Abstract (word count = 159)Small heat-shock proteins (sHsps) are a diverse family of intracellular molecular chaperone proteins that play a critical role in preventing protein unfolding, misfolding and aggregation, particularly under stress conditions such as elevated temperature, oxidation and infection. In doing so, they assist in the maintenance of protein homeostasis (proteostasis) and thereby avoid the deleterious effects that result from loss of protein function and/or protein aggregation. The chaperone properties of sHsps are therefore employed extensively in many tissues to prevent the development of diseases associated with protein aggregation. There has been much research into the structure and mechanism of chaperone action of sHsps over approximately the past 30 years, and significant progress has been made of late, however, there are still many unanswered questions relating to these aspects of sHsps. In this review, we outline some of the recent advances in understanding the structure and function of mammalian sHsps, particularly in the context of their many and varied roles in disease.

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“…Interestingly, the E164X mutant showed a red shift in the minimum indicating that the deletion of the second of the two glutamic acid residues in the REEK motif does introduce a detectable change in secondary structure. A recent study showed an R163X mutant with significantly altered secondary structure compared with the wild type (27). The frameshift mutation, 464delCT, introduced a small blue shift of the minimum to 208 nm and the appearance of a shoulder at ϳ223 nm, both of which are indicative of increased helicity.…”

Section: Expression and Purification Of The C-terminal Extensionmentioning

confidence: 99%

“…Stepwise removal of C-terminal sequences from ␣A-crystallin, including the REEK motif, not only reduced oligomerization, but also resulted in the sequential loss of chaperone activity (26), despite the fact that the IX(I/V) motif was retained in the mutants investigated. Mutations of lysine and glutamic acid residues in the C-terminal extension can improve chaperone activity against amyloid fibril formation providing a potential application of ␣B-crystallin in the treatment of amyloid-related diseases (27). In ␣B-crystallin, there has not been any systematic analysis of the role of the C-terminal extension and therefore no potential explanation of whether the three mutations (450delA, Q151X, and 464delCT) cause the various diseases via the loss of chaperone function or perhaps by a different mechanism.…”

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confidence: 99%

Truncation of αB-Crystallin by the Myopathy-causing Q151X Mutation Significantly Destabilizes the Protein Leading to Aggregate Formation in Transfected Cells

Hayes

Devlin

Quinlan

2008

Journal of Biological Chemistry

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Here we investigate the effects of a myopathy-causing mutation in ␣B-crystallin, Q151X, upon its structure and function. This mutation removes the C-terminal domain of ␣B-crystallin, which is expected to compromise both its oligomerization and chaperone activity. We compared this to two other ␣B-crystallin mutants (450delA, 464delCT) and also to a series of C-terminal truncations (E164X, E165X, K174X, and A171X). We find that the effects of the Q151X mutation were not always as predicted. Specifically, we have found that although the Q151X mutation decreased oligomerization of ␣B-crystallin and even increased some chaperone activities, it also significantly destabilized ␣B-crystallin causing it to self-aggregate. This conclusion was supported by our analyses of both the other disease-causing mutants and the series of C-terminal truncation constructs of ␣B-crystallin. The 450delA and 464delCT mutants could only be refolded and assayed as a complex with wild type ␣B-crystallin, which was not the case for Q151X ␣B-crystallin. From these studies, we conclude that all three disease-causing mutations (450delA, 464delCT, and Q151X) in the C-terminal extension destabilize ␣B-crystallin and increase its tendency to self-aggregate. We propose that it is this, rather than a catastrophic loss of chaperone activity, which is a major factor in the development of the reported diseases for the three disease-causing mutations studied here. In support of this hypothesis, we show that Q151X ␣B-crystallin is found mainly in the insoluble fraction of cell extracts from transient transfected cells, due to the formation of cytoplasmic aggregates.The crystallins were first identified as the major proteins of the eye lens and their subsequent classification into ␣-, ␤-, and ␥-crystallins (1) allowed different functional properties to be assigned to the ␣-and ␤␥-crystallin groups (2). The ␣-crystallins are two distinct proteins derived from two separate genes, ␣A-and ␣B-crystallin (2). Whereas ␣A-crystallin is almost entirely lens specific (3), ␣B-crystallin is expressed widely in other tissues, most notably astrocytes (4) and muscle (5). In the early 1980s, pioneering work from Klemenz and co-workers (6) and Horwitz (7) laboratories established that ␣B-crystallin was a protein chaperone and a member of the small heat shock protein (sHSP) 2 family, that is now known to comprise 10 different human proteins (8).When the first mutation (R120G) in ␣B-crystallin was reported, it was found to cause both cataract and desmin-related myopathy in the affected patients (9). Characteristic histopathological aggregates of the muscle intermediate filament protein desmin were observed (9), strongly suggesting that there is an important functional interaction between intermediate filaments and ␣B-crystallin. Subsequent analyses showed that the R120G ␣B-crystallin mutation induced increased binding for desmin intermediate filaments (10), providing an explanation for the formation of the desmin aggregates in the muscles of the affected individuals. Coinci...

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Site-Directed Mutations in the C-Terminal Extension of Human αB-Crystallin Affect Chaperone Function and Block Amyloid Fibril Formation

Cited by 42 publications

References 68 publications

Crystallin proteins and amyloid fibrils

Crystallin proteins and amyloid fibrils

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

Truncation of αB-Crystallin by the Myopathy-causing Q151X Mutation Significantly Destabilizes the Protein Leading to Aggregate Formation in Transfected Cells

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