Purification and characterization of the chaperone-like Hsp26 from Saccharomyces cerevisiae

Ferreira, R.; Andrade, Leonardo R.; Dutra, Marcio B; Souza, Marcos Farina de; Paschoalin, Vânia Margaret Flosi; Silva, Joab Trajano

doi:10.1016/j.pep.2006.02.006

Cited by 10 publications

(5 citation statements)

References 31 publications

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“…For example, some of the only outliers in the mild heat shock time course from [6] occur towards the end of a shift from 29 C to 33 C (Figure 5B). These include HSP26 and HSP78 , both known heat shock chaperones [25],[26]. Three genes of unknown function ( YLR327C , MOH1 and the neighboring dubious ORF YBL048W , and TMA10 ) are also outliers in this condition, which is evidence that these genes may have specific expression responses (and thus biological functions) during heat shock.…”

Section: Resultsmentioning

confidence: 89%

Predicting Cellular Growth from Gene Expression Signatures

et al. 2009

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Maintaining balanced growth in a changing environment is a fundamental systems-level challenge for cellular physiology, particularly in microorganisms. While the complete set of regulatory and functional pathways supporting growth and cellular proliferation are not yet known, portions of them are well understood. In particular, cellular proliferation is governed by mechanisms that are highly conserved from unicellular to multicellular organisms, and the disruption of these processes in metazoans is a major factor in the development of cancer. In this paper, we develop statistical methodology to identify quantitative aspects of the regulatory mechanisms underlying cellular proliferation in Saccharomyces cerevisiae. We find that the expression levels of a small set of genes can be exploited to predict the instantaneous growth rate of any cellular culture with high accuracy. The predictions obtained in this fashion are robust to changing biological conditions, experimental methods, and technological platforms. The proposed model is also effective in predicting growth rates for the related yeast Saccharomyces bayanus and the highly diverged yeast Schizosaccharomyces pombe, suggesting that the underlying regulatory signature is conserved across a wide range of unicellular evolution. We investigate the biological significance of the gene expression signature that the predictions are based upon from multiple perspectives: by perturbing the regulatory network through the Ras/PKA pathway, observing strong upregulation of growth rate even in the absence of appropriate nutrients, and discovering putative transcription factor binding sites, observing enrichment in growth-correlated genes. More broadly, the proposed methodology enables biological insights about growth at an instantaneous time scale, inaccessible by direct experimental methods. Data and tools enabling others to apply our methods are available at http://function.princeton.edu/growthrate.

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Section: Resultsmentioning

confidence: 89%

Predicting Cellular Growth from Gene Expression Signatures

et al. 2009

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“…Using both SEC-MALS and MS approaches, we have shown that at ambient temperature this protein exists as a polydisperse ensemble comprising numerous oligomeric states. Previous reports have suggested the presence of multiple oligomeric states for Sc HSP26 (Bentley et al, 1992; Ferreira et al, 2006), but here we have quantified their relative populations. Using a tandem MS technique, we have demonstrated the 24-mer to be “anomalously” abundant at 25°C, consistent with previous reports that have culminated in a cryo-electron microscopy-derived 3D structure of this oligomeric state (White et al, 2006).…”

Section: Discussionmentioning

confidence: 99%

The Quaternary Organization and Dynamics of the Molecular Chaperone HSP26 Are Thermally Regulated

Benesch¹,

Aquilina

Baldwin

et al. 2010

Chemistry & Biology

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SUMMARY The function of ScHSP26 is thermally controlled: the heat shock that causes the destabilization of target proteins leads to its activation as a molecular chaperone. We investigate the structural and dynamical properties of ScHSP26 oligomers through a combination of multiangle light scattering, fluorescence spectroscopy, NMR spectroscopy, and mass spec-trometry. We show that ScHSP26 exists as a heterogeneous oligomeric ensemble at room temperature. At heat-shock temperatures, two shifts in equilibria are observed: toward dissociation and to larger oligomers. We examine the quaternary dynamics of these oligomers by investigating the rate of exchange of subunits between them and find that this not only increases with temperature but proceeds via two separate processes. This is consistent with a conformational change of the oligomers at elevated temperatures which regulates the disassembly rates of this thermally activated protein.

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“…Chaperone activity for 1-Cys Prx6 proteins has not yet been reported. In order to enquire potential chaperone activity of AnPrx6, a chaperone assay that uses Hen Egg White Lysozyme (HEWL) as substrate was employed 60 . Lysozyme is a monomeric protein of 14.6 kDa with four disulphide bonds stabilizing its tertiary structure.…”

Section: Resultsmentioning

confidence: 99%

“…The assay is based on the thermal aggregation of lysozyme and follows the protocol established by Ferreira et al . 60 . We used 360 μl of an aqueous solution containing 50 mM NaP i buffer pH 7.1 and 30 mM DTT and placed it for 5 minutes in a water bath maintained at 43 °C.…”

Section: Methodsmentioning

confidence: 99%

Active-site plasticity revealed in the asymmetric dimer of AnPrx6 the 1-Cys peroxiredoxin and molecular chaperone from Anabaena sp. PCC 7120

Mishra

Hall

Locmelis

et al. 2017

Sci Rep

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Peroxiredoxins (Prxs) are vital regulators of intracellular reactive oxygen species levels in all living organisms. Their activity depends on one or two catalytically active cysteine residues, the peroxidatic Cys (CP) and, if present, the resolving Cys (CR). A detailed catalytic cycle has been derived for typical 2-Cys Prxs, however, little is known about the catalytic cycle of 1-Cys Prxs. We have characterized Prx6 from the cyanobacterium Anabaena sp. strain PCC7120 (AnPrx6) and found that in addition to the expected peroxidase activity, AnPrx6 can act as a molecular chaperone in its dimeric state, contrary to other Prxs. The AnPrx6 crystal structure at 2.3 Å resolution reveals different active site conformations in each monomer of the asymmetric obligate homo-dimer. Molecular dynamic simulations support the observed structural plasticity. A FSH motif, conserved in 1-Cys Prxs, precedes the active site PxxxTxxCp signature and might contribute to the 1-Cys Prx reaction cycle.

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Purification and characterization of the chaperone-like Hsp26 from Saccharomyces cerevisiae

Cited by 10 publications

References 31 publications

Predicting Cellular Growth from Gene Expression Signatures

Predicting Cellular Growth from Gene Expression Signatures

The Quaternary Organization and Dynamics of the Molecular Chaperone HSP26 Are Thermally Regulated

Active-site plasticity revealed in the asymmetric dimer of AnPrx6 the 1-Cys peroxiredoxin and molecular chaperone from Anabaena sp. PCC 7120

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