Stabilization of Proteins and Peptides in Diagnostic Immunological Assays by the Molecular Chaperone Hsp25

Ehrnsperger, Monika; Hergersberg, Christoph; Wienhues, Ulla; Nichtl, Alfons; Büchner, Johannes

doi:10.1006/abio.1998.2630

Cited by 46 publications

(32 citation statements)

References 37 publications

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“…Interestingly, tetramers, which we defined here as important intermediates in the assembly process of Hsp25, have also been shown to bind unfolded protein (22), indicating that different oligomeric Hsp25 species can posses chaperone activity (see Fig. 7).…”

Section: Discussionmentioning

confidence: 87%

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The Dynamics of Hsp25 Quaternary Structure

Ehrnsperger¹,

Lilie²,

Gaestel³

et al. 1999

Journal of Biological Chemistry

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155

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Small heat shock proteins (sHsps), including ␣-crystallin, represent a conserved and ubiquitous family of proteins. They form large oligomers, ranging in size from 140 to more than 800 kDa, which seem to be important for the interaction with non-native proteins as molecular chaperones. Here we analyzed the stability and oligomeric structure of murine Hsp25 and its correlation with function. Upon unfolding, the tertiary and quaternary structure of Hsp25 is rapidly lost, whereas the secondary structure remains remarkably stable. Unfolding is completely reversible, leading to native hexadecameric structures. These oligomers are in a concentration-dependent equilibrium with tetramers and dimers, indicating that tetramers assembled from dimers represent the basic building blocks of Hsp25 oligomers. At high temperatures, the Hsp25 complexes increase in molecular mass, consistent with the appearance of "heat shock granules" in vivo after heat treatment. This high molecular mass "heat shock form" of Hsp25 is in a slow equilibrium with hexadecameric Hsp25. Thus, it does not represent an off-pathway reaction. Interestingly, the heat shock form exhibits unchanged chaperone activity even after incubation at 80°C. We conclude that Hsp25 is a dynamic tetramer of tetramers with a unique ability to refold and reassemble into its active quaternary structure after denaturation. So-called heat shock granules, which have been reported to appear in response to stress, seem to represent a novel functional species of Hsp25.Small heat shock proteins (sHsps), 1 exhibiting a monomeric molecular mass of 9 -42 kDa, are expressed in all organisms investigated so far. Because of functional as well as structural homologies, ␣-crystallin, a major mammalian eye lens protein, which is also expressed in non-lenticular tissue, is a member of this protein family (1-4). Although the overall homology between different sHsps is rather low, they are grouped together based on conserved sequences in the C-terminal half of the protein and short, conserved, phenylalanine-rich stretches near the N terminus of the protein (5, 6). Mammalian sHsps are expressed constitutively even under physiological conditions. However, stress factors such as heat shock induce a strong up-regulation of protein levels by 10 -20-fold to maximum concentrations of 0.1% of the cellular protein (7, 8). Overexpression of different mammalian sHsps increases cellular thermoresistance significantly (9, 10). Furthermore, sHsps have been suggested to function in different, seemingly unrelated processes like RNA stabilization (11), interaction with the cytoskeleton (12, 13), or apoptosis (14). Interestingly, sHsps are also overexpressed in several cancers and neurodegenerative diseases like Alzheimer's disease or multiple sclerosis (15-17). In plants, five different classes of sHsps have been identified, which are partly localized in organella (8,18).In vitro sHsps act as molecular chaperones in preventing unfolded proteins from irreversible aggregation (3,4,19) and, in cooperation with o...

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Section: Discussionmentioning

confidence: 87%

“…Hsp70 and ATP, facilitate productive refolding of unfolded proteins (20,21). In this context, sHsps are more efficient than the model chaperone GroEL, due to their high binding capacity of up to one substrate molecule per sHsp subunit (21)(22)(23)(24).…”

mentioning

confidence: 99%

The Dynamics of Hsp25 Quaternary Structure

Ehrnsperger¹,

Lilie²,

Gaestel³

et al. 1999

Journal of Biological Chemistry

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155

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“…Unlike other ␣-Hsps, yeast Hsp26 requires temperature-assisted dissociation of a 24-mer into dimeric species in order to become an efficient chaperone (112). Tetramers of murine Hsp25 that occur as intermediates in the assembly of 16-mers have been shown to bind troponin T, a structurally labile marker protein for myocardial cell damage (73). This short list of observations already implies that the correlation between the oligomeric status and chaperone activity is different for individual ␣-Hsps.…”

Section: Homo-oligomeric Complexesmentioning

confidence: 99%

α-Crystallin-Type Heat Shock Proteins: Socializing Minichaperones in the Context of a Multichaperone Network

Narberhaus

2002

Microbiol Mol Biol Rev

498

415

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SUMMARY α-Crystallins were originally recognized as proteins contributing to the transparency of the mammalian eye lens. Subsequently, they have been found in many, but not all, members of the Archaea, Bacteria, and Eucarya. Most members of the diverse α-crystallin family have four common structural and functional features: (i) a small monomeric molecular mass between 12 and 43 kDa; (ii) the formation of large oligomeric complexes; (iii) the presence of a moderately conserved central region, the so-called α-crystallin domain; and (iv) molecular chaperone activity. Since α-crystallins are induced by a temperature upshift in many organisms, they are often referred to as small heat shock proteins (sHsps) or, more accurately, α-Hsps. α-Crystallins are integrated into a highly flexible and synergistic multichaperone network evolved to secure protein quality control in the cell. Their chaperone activity is limited to the binding of unfolding intermediates in order to protect them from irreversible aggregation. Productive release and refolding of captured proteins into the native state requires close cooperation with other cellular chaperones. In addition, α-Hsps seem to play an important role in membrane stabilization. The review compiles information on the abundance, sequence conservation, regulation, structure, and function of α-Hsps with an emphasis on the microbial members of this chaperone family.

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“…Thus, they act as molecular traps that protect the cell from the consequences of irreversible protein aggregation. This holdase function of sHsps enables to separate the binding of unfolded polypeptides from the refolding process thereby sustaining protein homeostasis (4,(7)(8)(9).…”

mentioning

confidence: 99%

The eye lens chaperone α-crystallin forms defined globular assemblies

Peschek

Braun

Franzmann

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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127

121

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␣-Crystallins are molecular chaperones that protect vertebrate eye lens proteins from detrimental protein aggregation. ␣B-Crystallin, 1 of the 2 ␣-crystallin isoforms, is also associated with myopathies and neuropathological diseases. Despite the importance of ␣-crystallins in protein homeostasis, only little is known about their quaternary structures because of their seemingly polydisperse nature. Here, we analyzed the structures of recombinant ␣-crystallins using biophysical methods. In contrast to previous reports, we show that ␣B-crystallin assembles into defined oligomers consisting of 24 subunits. The 3-dimensional (3D) reconstruction of ␣B-crystallin by electron microscopy reveals a spherelike structure with large openings to the interior of the protein.␣A-Crystallin forms, in addition to complexes of 24 subunits, also smaller oligomers and large clusters consisting of individual oligomers. This propensity might explain the previously reported polydisperse nature of ␣-crystallin.electron microscopy ͉ molecular chaperone ͉ protein aggregation ͉ small heat shock protein ͉ stress response

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Stabilization of Proteins and Peptides in Diagnostic Immunological Assays by the Molecular Chaperone Hsp25

Cited by 46 publications

References 37 publications

The Dynamics of Hsp25 Quaternary Structure

The Dynamics of Hsp25 Quaternary Structure

α-Crystallin-Type Heat Shock Proteins: Socializing Minichaperones in the Context of a Multichaperone Network

The eye lens chaperone α-crystallin forms defined globular assemblies

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