scholarly journals Mammalian Hsp22 is a heat-inducible small heat-shock protein with chaperone-like activity

2004 ◽  
Vol 381 (2) ◽  
pp. 379-387 ◽  
Author(s):  
Tirumala Kumar CHOWDARY ◽  
Bakthisaran RAMAN ◽  
Tangirala RAMAKRISHNA ◽  
Chintalagiri Mohan RAO
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Quaternary Structure ◽  
Citrate Synthase ◽  
Gradient Centrifugation ◽  
Gel Filtration ◽  
Nucleotide Sequence Analysis ◽  
Heat Shock Factor 1 ◽  
Induced Aggregation

A newly identified 22 kDa protein that interacts with Hsp27 (heat-shock protein 27) was shown to possess the characteristic α-crystallin domain, hence named Hsp22, and categorized as a member of the sHsp (small Hsp) family. Independent studies from different laboratories reported the protein with different names such as Hsp22, H11 kinase, E2IG1 and HspB8. We have identified, on the basis of the nucleotide sequence analysis, putative heat-shock factor 1 binding sites upstream of the Hsp22 translation start site. We demonstrate that indeed Hsp22 is heat-inducible. We show, in vitro, chaperone-like activity of Hsp22 in preventing dithiothreitol-induced aggregation of insulin and thermal aggregation of citrate synthase. We have cloned rat Hsp22, overexpressed and purified the protein to homogeneity and studied its structural and functional aspects. We find that Hsp22 fragments on storage. MS analysis of fragments suggests that the fragmentation might be due to the presence of labile peptide bonds. We have established conditions to improve its stability. Far-UV CD indicates a randomly coiled structure for Hsp22. Quaternary structure analyses by glycerol density-gradient centrifugation and gel filtration chromatography show that Hsp22 exists as a monomer in vitro, unlike other members of the sHsp family. Hsp22 exhibits significantly exposed hydrophobic surfaces as reported by bis-8-anilinonaphthalene-l-sulphonic acid fluorescence. We find that the chaperone-like activity is temperature dependent. Thus Hsp22 appears to be a true member of the sHsp family, which exists as a monomer in vitro and exhibits chaperone-like activity.

ArHsp21, a developmentally regulated small heat-shock protein synthesized in diapausing embryos of Artemia franciscana

2008 ◽  
Vol 411 (3) ◽  
pp. 605-611 ◽  
Author(s):  
Zhijun Qiu ◽  
Thomas H. MacRae
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Citrate Synthase ◽  
Sequence Similarity ◽  
Single Gene ◽  
Small Heat Shock Protein ◽  
Subtractive Hybridization ◽  
Artemia Franciscana ◽  
Induced Aggregation

Embryos of the crustacean, Artemia franciscana, undergo alternative developmental pathways, producing either larvae or encysted embryos (cysts). The cysts enter diapause, characterized by exceptionally high resistance to environmental stress, a condition thought to involve the sHSP (small heat-shock protein), p26. Subtractive hybridization has revealed another sHSP, termed ArHsp21, in diapause-destined Artemia embryos. ArHsp21 shares sequence similarity with p26 and sHSPs from other organisms, especially in the α-crystallin domain. ArHsp21 is the product of a single gene and its synthesis occurred exclusively in diapause-destined embryos. Specifically, ArHsp21 mRNA appeared 2 days post-fertilization, followed 1 day later by the protein, and then increased until embryo release at day 5. No ArHsp21 protein was detected in embryos developing directly into larvae, although there was a small amount of mRNA at 3 days post-fertilization. The protein was degraded during post-diapause development and had disappeared completely from second instar larvae. ArHsp21 formed large oligomers in encysted embryos and transformed bacteria. When purified from bacteria, ArHsp21 functioned as a molecular chaperone in vitro, preventing heat-induced aggregation of citrate synthase and reduction-driven denaturation of insulin. Sequence characteristics, synthesis patterns and functional properties demonstrate clearly that ArHsp21 is an sHSP able to chaperone other proteins and contribute to stress tolerance during diapause. As such, ArHsp21 would augment p26 chaperone activity and it may also possess novel activities that benefit Artemia embryos exposed to stress.

scholarly journals Changes in the quaternary structure and function of MjHSP16.5 attributable to deletion of the IXI motif and introduction of the substitution, R107G, in the α -crystallin domain

2013 ◽  
Vol 368 (1617) ◽  
pp. 20120327 ◽  
Author(s):  
Roy A. Quinlan ◽  
Yan Zhang ◽  
Andrew Lansbury ◽  
Ian Williamson ◽  
Ehmke Pohl ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Room Temperature ◽  
Quaternary Structure ◽  
Citrate Synthase ◽  
Small Heat Shock Protein ◽  
Octahedral Symmetry ◽  
Functional Consequences ◽  
And Function ◽  
Induced Aggregation

The archael small heat-shock protein (sHSP), MjHSP16.5, forms a 24-subunit oligomer with octahedral symmetry. Here, we demonstrate that the IXI motif present in the C-terminal domain is necessary for the oligomerization of MjHSP16.5. Removal increased the in vitro chaperone activity with citrate synthase as the client protein. Less predictable were the effects of the R107G substitution in MjHSP16.5 because of the differences in the oligomerization of metazoan and non-metazoan sHSPs. We present the crystal structure for MjHSP16.5 R107G and compare this with an improved (2.5 Å) crystal structure for wild-type (WT) MjHSP16.5. Although no significant structural differences were found in the crystal, using cryo-electron microscopy, we identified two 24mer species with octahedral symmetry for the WT MjHSP16.5 both at room temperature and at 60°C, all showing two major species with the same diameter of 12.4 nm. Similarly, at room temperature, there are also two kinds of 12.4 nm oligomers for R107G MjHSP16.5, but in the 60°C sample, a larger 24mer species with a diameter of 13.6 nm was observed with significant changes in the fourfold symmetry axis and dimer–dimer interface. This highly conserved arginine, therefore, contributes to the quaternary organization of non-metazoan sHSP oligomers. Potentially, the R107G substitution has functional consequences as R107G MjHSP16.5 was far superior to the WT protein in protecting β L -crystallin against heat-induced aggregation.

The reassembling process of the nonameric Mycobacterium tuberculosis small heat-shock protein Hsp16.3 occurs via a stepwise mechanism

2002 ◽  
Vol 363 (2) ◽  
pp. 329-334 ◽  
Author(s):  
Xiuguang FENG ◽  
Sufang HUANG ◽  
Xinmiao FU ◽  
Abuduaini ABULIMITI ◽  
Zengyi CHANG
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Quaternary Structure ◽  
Small Heat Shock Protein ◽  
Oligomeric Proteins ◽  
Stepwise Mechanism ◽  
Pore Gradient ◽  
Modified Urea

Conditions are reported under which the reassembled intermediates of the heat-shock protein Hsp16.3 after being denatured in 8M urea were detected by mainly using urea-gradient PAGE (with modifications) and urea-denaturing pore-gradient PAGE. Hsp16.3 is the small heat-shock protein from Mycobacterium tuberculosis, which exists as a specific nonamer and was proposed to form a trimer-of-trimers structure. The refolding and reassembling of this protein was achieved rapidly by dilution or dialysis, suggesting an effectively spontaneous recovery of quaternary structure. Data presented in this report demonstrate that the in vitro reassembling process of Hsp16.3 protein occurs through a spontaneous and effective stepwise mechanism. Modified urea-gradient PAGE may provide a general method for studying the reassembling processes of other oligomeric proteins.

scholarly journals Heat Shock Protein 70 Improves In Vitro Embryo Yield and Quality from Heat Stressed Bovine Oocytes

Animals ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 1794
Author(s):  
Konstantina Stamperna ◽  
Themistoklis Giannoulis ◽  
Eleni Dovolou ◽  
Maria Kalemkeridou ◽  
Ioannis Nanas ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Heat Shock Protein 70 ◽  
Cumulus Cells ◽  
Intramolecular Interactions ◽  
Developmental Competence ◽  
Embryo Yield ◽  
Bovine Oocytes

Heat shock protein 70 (HSP70) is a chaperon that stabilizes unfolded or partially folded proteins, preventing inappropriate inter- and intramolecular interactions. Here, we examined the developmental competence of in vitro matured oocytes exposed to heat stress with or without HSP70. Bovine oocytes were matured for 24 h at 39 °C without (group C39) or with HSP70 (group H39) and at 41 °C for the first 6 h, followed by 16 h at 39 °C with (group H41) or without HSP70 (group C41). After insemination, zygotes were cultured for 9 days at 39 °C. Cleavage and embryo yield were assessed 48 h post insemination and on days 7, 8, 9, respectively. Gene expression was assessed by RT-PCR in oocytes, cumulus cells and blastocysts. In C41, blastocysts formation rate was lower than in C39 and on day 9 it was lower than in H41. In oocytes, HSP70 enhanced the expression of three HSP genes regardless of incubation temperature. HSP70 at 39 °C led to tight coordination of gene expression in oocytes and blastocysts, but not in cumulus cells. Our results imply that HSP70, by preventing apoptosis, supporting signal transduction, and increasing antioxidant protection of the embryo, protects heat stressed maturing bovine oocyte and restores its developmental competence.

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