The apparent molecular size of native α‐crystallin B in non‐lenticular tissues

Chiesa, Raúl; McDermott, Martin J.; Mann, Eric; Spector, Abraham

doi:10.1016/0014-5793(90)81013-e

Cited by 28 publications

(9 citation statements)

References 23 publications

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“…It is possible that aA and aB exist as separate inultimeric forms of similar mass. While this work was in progress, Chiesa et al [42] reported that aB in the extralenticular tissues was an aggregate of about 800 kDa molecular mass, the same as reported earlier for a-crystallin (containing aA and a s ) in the lens by the same laboratory [4]. These investigators did not analyze purified aB to allow a direct assessment of the discrepancy between their and our determinations.…”

Section: Discussionsupporting

confidence: 55%

αB‐crystallin exists as an independent protein in the heart and in the lens

Bhat

Horwitz

Srinivasan

et al. 1991

European Journal of Biochemistry

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-EJB 91 0543 aB-crystallin, a polypeptide of molecular mass 22 kDa, is considered to be one of two subunits (aA and aB) of the multimeric lens-specific protein, a-crystallin. Recent demonstrations of the extralenticular presence of aB-crystallin have suggested that outside of the lens, this polypeptide may have functions independent of aA. Within the lens however, as part of the protein a-crystallin, its function is assumed to be structural. In an effort to investigate the functional status of aB-crystallin in the lens, we have characterized this polypeptide in the rat heart and the human lens.Unequivocal identity of aB-crystallin in the rat heart and the rat lens was established by the sequence analyses of the respective cDNA clones. Size exclusion chromatography (FPLC) and immunoblotting showed that in the rat heart, aB-crystallin exists as an aggregate of 300-400 kDa average molecular mass, similar to that of purified ixB-crystallin isolated from bovine lens. Interestingly, analysis of the human lens proteins by iinmunoblotting showed that, with age, unlike aAcrystallin, the aB subunit remains detectable in the soluble fractions derived from normal lenses as old as 82 years. Importantly, the average molecular mass of the aB subunit in the soluble fractions prepared from 60-80-year-old human lens nuclei was also found to be 300-400 kDa. These data lead to the conclusion that aB-crystallin may exist as an independent protein not only in non-lens tissues (e.g. heart) but in the lens as well a-Crystallin is one of the predominant multimeric structural proteins of the mammalian lens, known to be composed of two primary gene products, aA and aB [l, 21. It is the first crystallin to appear during mammalian lens development and is known to undergo a number of gerontological changes in its structure [3 -71. Its concentration in the lens reaches up to 30% of the total soluble protein and is, therefore, one of the chief contributory factors for the maintenance of transparency [8], the main functional attribute of the ocular lens.a-Crystallin is involved in the formation of high-molecular-mass protein aggregates during aging and is eventually lost from the soluble pool of the adult lens proteins. It is also one of the main components of very-high-molecular-mass insoluble aggregates present in opaque lenses [7, 9-121. The two polypeptides (aA and a s ) have been historically treated as the two integral subunits of one structural protein, cc-crystallin [l]. However, most of the studies conducted with acrystallin have been restricted to aA because of its predominance in the lens (aA: aB, 3 : 1) [I]. Observations made primarily with aA have been extrapolated to aB, including the presumed structural function for this polypeptide [13]. This tentative status has been further fortified by the fact that a A and aB share 57% sequence similarity at the polypeptide level [14].Both of these proteins also share significant sequence similarity with small heat-shock proteins and are, therefore, re- garded as members of the small heat-...

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Section: Discussionsupporting

confidence: 55%

αB‐crystallin exists as an independent protein in the heart and in the lens

Bhat

Horwitz

Srinivasan

et al. 1991

European Journal of Biochemistry

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“…Under normal conditions, small Hsps generally form large oligomers up to ~0.8 MDa in size [19][20][21][22]. There seem to be variations in the apparent stability of the oligomers.…”

Section: Small Hsps Form Large Dynamic Oligomersmentioning

confidence: 99%

The small heat shock proteins and their clients

Nakamoto

Vı́gh

2006

Cell. Mol. Life Sci.

273

218

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Small heat shock proteins are ubiquitous proteins found throughout all kingdoms. One of the most notable features is their large oligomeric structures with conserved structural organization. It is well documented that small heat shock proteins can capture unfolding proteins to form stable complexes and prevent their irreversible aggregation. In addition, small heat shock proteins coaggregate with aggregation-prone proteins for subsequent, efficient disaggregation of the protein aggregates. The release of substrate proteins from the transient reservoirs, i.e. complexes and aggregates with small heat shock proteins, and their refolding require cooperation with ATP-dependent chaperone systems. The amphitropic small heat shock proteins were shown to associate with membranes, although they do not contain transmembrane domains or signal sequences. Recent studies indicate that small heat shock proteins play an important role in membrane quality control and thereby potentially contribute to the maintenance of membrane integrity especially under stress conditions.

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“…The oligomeric size of differently phosphorylated Hsp27 was analyzed by size exclusion liquid chromatography using Superose 6. As determined by a number of different methods (12,13,23,38,(42)(43)(44), nonphosphorylated sHsps form complexes of an average molecular mass of 200 -800 kDa indicating a complex of 12 to nearly 40 sHsp monomers. Our analysis shows that nonphosphorylated Hsp27 exhibits an average mass of 530 kDa which correlates with an oligomer of about 24 subunits (Fig.…”

Section: Phosphorylation-regulated Oligomerization Of Hsp27 Inmentioning

confidence: 99%

Regulation of Hsp27 Oligomerization, Chaperone Function, and Protective Activity against Oxidative Stress/Tumor Necrosis Factor α by Phosphorylation

Rogalla

Ehrnsperger

Préville

et al. 1999

Journal of Biological Chemistry

690

663

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The small heat shock proteins (sHsps) from human (Hsp27) and mouse (Hsp25) form large oligomers which can act as molecular chaperones in vitro and protect cells from heat shock and oxidative stress when overexpressed. In addition, mammalian sHsps are rapidly phosphorylated by MAPKAP kinase 2/3 at two or three serine residues in response to various extracellular stresses. Here we analyze the effect of sHsp phosphorylation on its quaternary structure, chaperone function, and protection against oxidative stress. We show that in vitro phosphorylation of recombinant sHsp as well as molecular mimicry of Hsp27 phosphorylation lead to a significant decrease of the oligomeric size. We demonstrate that both phosphorylated sHsps and the triple mutant Hsp27-S15D,S78D,S82D show significantly decreased abilities to act as molecular chaperones suppressing thermal denaturation and facilitating refolding of citrate synthase in vitro. In parallel, Hsp27 and its mutants were analyzed for their ability to confer resistance against oxidative stress when overexpressed in L929 and 13.S.1.24 cells. While wild type Hsp27 confers resistance, the triple mutant S15D,S78D,S82D cannot protect against oxidative stress effectively. These data indicate that large oligomers of sHsps are necessary for chaperone action and resistance against oxidative stress whereas phosphorylation down-regulates these activities by dissociation of sHsp complexes to tetramers. Small heat shock proteins (sHsps)1 are constitutively expressed in virtually all organisms and exhibit a monomeric molecular mass of 15-42 kDa (for a recent review see Ref. 1). Within the cell they can form oligomeric complexes of up to 1 MDa (2). Overexpression of different mammalian sHsps increases cellular thermoresistance to a significant degree (3, 4). sHsps can, furthermore, function in different, seemingly unrelated processes like RNA stabilization (5), interaction with the cytoskeleton (6, 7), or apoptosis (8, 9). In vitro sHsps act as molecular chaperones preventing unfolded proteins from irreversible aggregation (10 -12) and, in cooperation with other factors, e.g. Hsp70 and ATP, facilitating productive refolding of unfolded proteins (13,14).In mammalian cells certain sHsps, e.g. mouse Hsp25 or human Hsp27, form a converging element of the cellular stress response since they show both a stress-induced increase in expression and phosphorylation. Under heat shock conditions increased phosphorylation can be detected after several minutes while changes in expression are detected after several hours (15). The rapid stress-induced phosphorylation is the result of stimulation of the p38 MAP kinase cascade and subsequent activation of MAPKAP kinases 2 and 3 which directly phosphorylate mammalian sHsps (16, 17) at several distinct sites (18,19). Since sHsp phosphorylation and stress-induced expression show different kinetics, it is assumed that phosphorylation of the pre-existing constitutively expressed sHsps is a first phase of the stress response while the elevated expression at a time w...

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The apparent molecular size of native α‐crystallin B in non‐lenticular tissues

Cited by 28 publications

References 23 publications

αB‐crystallin exists as an independent protein in the heart and in the lens

αB‐crystallin exists as an independent protein in the heart and in the lens

The small heat shock proteins and their clients

Regulation of Hsp27 Oligomerization, Chaperone Function, and Protective Activity against Oxidative Stress/Tumor Necrosis Factor α by Phosphorylation

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