Cloning and characterization of a thermostable catfish αB-crystallin with chaperone-like activity at high temperatures

Yu, Chung-Ming; Chang, Gu-Gang; Chang, Hui-Chuan; Chiou, Shyh‐Horng

doi:10.1016/j.exer.2004.04.006

Cited by 11 publications

(10 citation statements)

References 55 publications

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“…However, the sequence homology between H. pylori AhpC and mammalian (human or mouse) peroxiredoxins is as high as 43%. On the other hand, the phylogenetic tree analysis similar to our previous report (29) showed that all of the three H. pylori AhpC proteins are more homologous to mammalian peroxiredoxins than to other bacterial AhpC proteins (Fig. 1C).…”

Section: Resultssupporting

confidence: 66%

“…The insulin substrate stock solution consisted of 8.5 mg of insulin dissolved in 1 ml of 0.1 M NaOH, 1.7 ml of 0.5 M NaH 2 PO 4 ͞Na 2 HPO 4 buffer (pH 6.8), and 1.7 ml of 1 M NaCl, and the total volume was adjusted to 17 ml with distilled H 2 O. The AhpC-dependent chaperone assay was carried out at 25°C by recording the turbidity change of OD 360 within 42 min upon the initiation of DTT-induced insulin aggregation or until the turbidity curve reaches a plateau (29).…”

mentioning

confidence: 99%

“…Each assay was performed by using 20 M each AhpC in a total volume of 1.0 ml of 50 mM potassium phosphate buffer (pH 7.0)͞0.1 M ammonium sulfate͞0.5 mM EDTA containing Trx system (5 M thioredoxin, 0.5 M thioredoxin reductase, and 150 M NADPH) and 1 mM H 2 O 2 as described (21). Chaperone-like activity of recombinant AhpC was studied based on the DTT-induced insulin reductive unfolding and chaperone-assisted refolding (29,41). The reaction mixture was prepared in a total volume of 1.0 ml of PBS buffer containing insulin and DTT.…”

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confidence: 99%

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The antioxidant protein alkylhydroperoxide reductase of Helicobacter pylori switches from a peroxide reductase to a molecular chaperone function

Chuang

Wu²,

Lo³

et al. 2006

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

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105

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Helicobacter pylori, an oxygen-sensitive microaerophilic bacterium, contains many antioxidant proteins, among which alkylhydroperoxide reductase (AhpC) is the most abundant. The function of AhpC is to protect H. pylori from a hyperoxidative environment by reduction of toxic organic hydroperoxides. We have found that the sequence of AhpC from H. pylori is more homologous to mammalian peroxiredoxins than to eubacterial AhpC. We have also found that the protein structure of AhpC could shift from lowmolecular-weight oligomers with peroxide-reductase activity to high-molecular-weight complexes with molecular-chaperone function under oxidative stresses. Time-course study by following the quaternary structural change of AhpC in vivo revealed that this enzyme changes from low-molecular-weight oligomers under normal microaerobic conditions or short-term oxidative shock to high-molecular-weight complexes after severe long-term oxidative stress. This study revealed that AhpC of H. pylori acts as a peroxide reductase in reducing organic hydroperoxides and as a molecular chaperone for prevention of protein misfolding under oxidative stress.peroxiredoxin ͉ oxidative stress ͉ phylogenetic comparison ͉ dual functionality ͉ quaternary structural change

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Section: Resultssupporting

confidence: 66%

mentioning

confidence: 99%

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confidence: 99%

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The antioxidant protein alkylhydroperoxide reductase of Helicobacter pylori switches from a peroxide reductase to a molecular chaperone function

Chuang

Wu²,

Lo³

et al. 2006

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

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132

105

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“…These findings indicate that calreticulin is a thermostable protein behaving similarly to small heat-shock protein family members like catfish ␣B-crystallin and HSP16.5 from Methanococcus jannaschii with reported secondary structure stabilities up to 60 -80°C (35,36). In addition to its secondary structural stability, several structural and functional features of calreticulin resemble those described for small heat shock protein family members (reviewed in Ref.…”

Section: Discussionmentioning

confidence: 67%

Calreticulin Is a Thermostable Protein with Distinct Structural Responses to Different Divalent Cation Environments

Wijeyesakere¹,

Gafni

Raghavan³

2011

Journal of Biological Chemistry

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Calreticulin is a soluble calcium-binding chaperone of the endoplasmic reticulum (ER) that is also detected on the cell surface and in the cytosol. Calreticulin contains a single high affinity calcium-binding site within a globular domain and multiple low affinity sites within a C-terminal acidic region. We show that the secondary structure of calreticulin is remarkably thermostable at a given calcium concentration. Rather than corresponding to complete unfolding events, heat-induced structural transitions observed for calreticulin relate to tertiary structural changes that expose hydrophobic residues and reduce protein rigidity. The thermostability and the overall secondary structure content of calreticulin are impacted by the divalent cation environment, with the ER range of calcium concentrations enhancing stability, and calciumdepleting or high calcium environments reducing stability. Furthermore, magnesium competes with calcium for binding to calreticulin and reduces thermostability. The acidic domain of calreticulin is an important mediator of calcium-dependent changes in secondary structure content and thermostability. Together, these studies indicate interactions between the globular and acidic domains of calreticulin that are impacted by divalent cations. These interactions influence the structure and stability of calreticulin, and are likely to determine the multiple functional activities of calreticulin in different subcellular environments.

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“…In teleost fish lens, the γ crystallins are the major heat-labile constituents (Fig.·2, Table·2). Sequence similarity between fish and mammalian α crystallins indicated that the former are also sHSPs and would have chaperone-like activity (Posner et al, 1999;Runkle et al, 2002), which has indeed shown to be so for recombinant αA crystallin from zebrafish in chaperone assays with DTT-denatured lysozyme (Posner, 2003), and for recombinant αB from carp with DTT-denatured insulin (Yu et al, 2004). Here we demonstrate that native α crystallin isolated from the lens of two other fish, Antarctic toothfish and bigeye tuna, are also effective molecular chaperones, with the ability to protect the same species heat-labile γ crystallins from aggregation induced by heat ( Fig.·5A-C), and chemical denaturation of non-lens protein (lysozyme) by DLdithiothreitol ( Fig.·5D-F).…”

Section: Discussionmentioning

confidence: 99%

Cold-stable eye lens crystallins of the Antarctic nototheniid toothfishDissostichus mawsoniNorman

Kiss

Mirarefi

Ramakrishnan

et al. 2004

Journal of Experimental Biology

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SUMMARY The eye lenses of the Antarctic nototheniid fishes that inhabit the perennially freezing Antarctic seawater are transparent at –2°C,whereas the cold-sensitive mammalian and tropical fish lenses display cold-induced cataract at 20°C and 7°C, respectively. No cold-cataract occurs in the giant Antarctic toothfish Dissostichus mawsoni lens when cooled to temperatures as low as –12°C, indicating highly cold-stable lens proteins. To investigate this cold stability, we characterised the lens crystallin proteins of the Antarctic toothfish, in parallel with those of the sub-tropical bigeye tuna Thunnus obesusand the endothermic cow Bos taurus, representing three disparate thermal climes (–2°C, 18°C and 37°C, respectively). Sizing chromatography resolved their lens crystallins into three groups,α/βH, β and γ, with γ crystallins being the most abundant (>40%) lens proteins in fish, in contrast to the cow lens where they comprise only 19%. The upper thermal stability of these crystallin components correlated with the body temperature of the species. In vitro chaperone assays showed that fish α crystallin can protect same-species γ crystallins from heat denaturation, as well as lysozyme from DTT-induced unfolding, and therefore are small Heat Shock Proteins (sHSP)like their mammalian counterparts. Dynamic light scattering measured an increase in size of αγ crystallin mixtures upon heating, which supports formation of the αγ complex as an integral part of the chaperone process. Surprisingly, in cross-species chaperone assays, tunaα crystallins only partly protected toothfish γ crystallins, while cow α crystallins completely failed to protect, indicating partial and no αγ interaction, respectively. Toothfish γ was likely to be the component that failed to interact, as the supernatant from a cowα plus toothfish γ incubation could chaperone cow γcrystallins in a subsequent heat incubation, indicating the presence of uncomplexed cow α. This suggests that the inability of toothfish γcrystallins to fully complex with tuna α, and not at all with the cowα crystallins, may have its basis in adaptive changes in the protein that relate to the extreme cold-stability of the toothfish lens.

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Cloning and characterization of a thermostable catfish αB-crystallin with chaperone-like activity at high temperatures

Cited by 11 publications

References 55 publications

The antioxidant protein alkylhydroperoxide reductase of Helicobacter pylori switches from a peroxide reductase to a molecular chaperone function

The antioxidant protein alkylhydroperoxide reductase of Helicobacter pylori switches from a peroxide reductase to a molecular chaperone function

Calreticulin Is a Thermostable Protein with Distinct Structural Responses to Different Divalent Cation Environments

Cold-stable eye lens crystallins of the Antarctic nototheniid toothfishDissostichus mawsoniNorman

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