In Vivo Substrate Diversity and Preference of Small Heat Shock Protein IbpB as Revealed by Using a Genetically Incorporated Photo-cross-linker

Fu, Xinmiao; Shi, Xiaodong; Yan, Linxuan; Zhang, Hanlin; Chang, Zengyi

doi:10.1074/jbc.m113.501817

Cited by 54 publications

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References 65 publications

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“…Uniquely, the sHSPs of both prokaryotic and eukaryotic sources have also been found to be associated with cell membranes (34)(35)(36)(37)(38). Over the years, we have tried to delineate the function and mechanism of sHSPs, from largely bacterial sources, via both in vitro and in vivo studies (39)(40)(41)(42)(43)(44)(45). To study the function and mechanism of sHSPs in C. elegans, we first focused on CeHSP17, which remains the least characterized such proteins in terms of function and properties among the small heat shock proteins of C. elegans (46) and is predicted by our bioinformatics analysis to be putatively localized in mitochondria (Table 1 ).…”

Section: Resultsmentioning

confidence: 99%

A Small Heat Shock Protein Enables Escherichia coli To Grow at a Lethal Temperature of 50 C Conceivably by Maintaining Cell Envelope Integrity

Ezemaduka

Shi

et al. 2014

Journal of Bacteriology

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It is essential for organisms to adapt to fluctuating growth temperatures. Escherichia coli, a model bacterium commonly used in research and industry, has been reported to grow at a temperature lower than 46.5°C. Here we report that the heterologous expression of the 17-kDa small heat shock protein from the nematode Caenorhabditis elegans, CeHSP17, enables E. coli cells to grow at 50°C, which is their highest growth temperature ever reported. Strikingly, CeHSP17 also rescues the thermal lethality of an E. coli mutant deficient in degP, which encodes a protein quality control factor localized in the periplasmic space. Mechanistically, we show that CeHSP17 is partially localized in the periplasmic space and associated with the inner membrane of E. coli, and it helps to maintain the cell envelope integrity of the E. coli cells at the lethal temperatures. Together, our data indicate that maintaining the cell envelope integrity is crucial for the E. coli cells to grow at high temperatures and also shed new light on the development of thermophilic bacteria for industrial application.T emperature is considered the most important single environmental factor that profoundly affects the structure and function of biomolecules. Each organism in nature has evolved to live at a certain optimal temperature range. Nonetheless, effective mechanisms have also been evolved for organisms to survive under nonoptimal temperature conditions, typically termed heat shock response (1, 2) and cold shock response (3). It is of great value to understand such mechanisms of living organisms for both unveiling the nature of life and exploring biotechnological application.Escherichia coli, being the most extensively studied bacterium and also a popularly utilized host cell for producing pharmaceutically important recombinant proteins, is known to be unable to grow at a temperature higher than 46.5°C (4-6). It has been widely reported that heterologous overexpression of certain exogenous molecular chaperones (7-11) or an endogenous transcriptional regulator (12) is able to significantly increase the viability of E. coli cells undergoing heat shock treatment at lethal temperatures (around 50°C). It is also well established that preincubating E. coli cells (13,14) and other organisms (reviewed in reference 15) at a sublethal temperature (e.g., 42°C) significantly increases the thermotolerance of the treated organisms at lethal temperatures. However, all these alternations usually would not allow the modified cells to permanently survive and even grow under such lethal temperatures. In two attempts at selecting heat-resistant phenotypes by using extensive experimental evolution, E. coli mutant strains that are able to grow at up to 48°C (16) or 48.5°C (5) were obtained, with growth at the latter temperature being partially related to the high level of expression of the molecular chaperone GroEL/GroES. In contrast, thermophilic bacteria have been found to grow effectively at optimal temperatures much higher than 50°C (17)(18)(19)(20). They are known...

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Section: Resultsmentioning

confidence: 99%

A Small Heat Shock Protein Enables Escherichia coli To Grow at a Lethal Temperature of 50 C Conceivably by Maintaining Cell Envelope Integrity

Ezemaduka

Shi

et al. 2014

Journal of Bacteriology

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“…In addition, a total of 94 proteins in the cell extract of bacterium Deinococcus radiodurans were found to co-aggregate with Hsp20.2 during thermal treatment [56]. Importantly, we have recently identified a total of 113 cellular proteins interacting with IbpB in living E. coli cells by using in vivo photo-crosslinking [54]. Retrospectively, these natural substrate proteins of specific sHSPs appear to have a variety of cellular functions, such as metabolism (e.g.…”

Section: Functional Diversity Of the Substrate Proteins Of Shspsmentioning

confidence: 99%

“…To clarify this, database searching was performed for three representative human sHSPs regarding their interacting partners. The results presented in Table 1 indicated that aB-crystallin binds a number of cellular proteins, including Hsp16.6-bound proteins after heat shock were co-purified, with 13 being identified [81]; IbpA-bound proteins were co-purified, with 42 being identified [82]; IbpB-bound proteins were covalently photo-crosslinked with IbpB in living cells and then co-purified, with 113 being identified [54]; Hsp20.2-bound proteins were co-purified from the thermally treated mixture of Hsp20.2 and cell extract, with 94 being identified [56]. The information for mammalian sHSPs-bound proteins was extracted from the protein-protein interaction databases (a uniform entry is provided at http:// www.ebi.ac.uk/Tools/webservices/psicquic/view/main.xhtml).…”

Section: Functional Diversity Of the Substrate Proteins Of Shspsmentioning

confidence: 99%

“…As such, sHSPs could act as robust chaperones under fluctuating conditions and enhance their chaperone activities at elevating temperatures. Importantly, accumulative evidence indicate that sHSPs also preferentially protect certain classes of functional proteins, such as translation-related proteins and metabolic enzymes [54,[56][57][58]. In this review, I will focus on how an sHSP activates its numerous multi-type substrate-binding residues and what functional proteins it protects in cells.…”

Section: Introductionmentioning

confidence: 99%

“…Recently, we have utilized the site-specific in vivo photo-crosslinking to probe the chaperone function and mechanism of sHSPs in Escherichia coli [54,55]. These studies suggested that prokaryotic sHSPs utilize numerous multi-type substrate-binding residues and hierarchically activate them in a temperature-dependent manner to capture a wide range of proteins.…”

Section: Introductionmentioning

confidence: 99%

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Chaperone function and mechanism of small heat-shock proteins

Fu¹

2014

ABBS

Self Cite

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Small heat-shock proteins (sHSPs) are ubiquitous ATP-independent molecular chaperones that play crucial roles in protein quality control in cells. They are able to prevent the aggregation and/or inactivation of various non-native substrate proteins and assist the refolding of these substrates independently or under the help of other ATP-dependent chaperones. Substrate recognition and binding by sHSPs are essential for their chaperone functions. This review focuses on what natural substrate proteins an sHSP protects and how it binds the substrates in cells under fluctuating conditions. It appears that sHSPs of prokaryotes, although being able to bind a wide range of cellular proteins, preferentially protect certain classes of functional proteins, such as translation-related proteins and metabolic enzymes, which may well explain why they could increase the resistance of host cells against various stresses. Mechanistically, the sHSPs of prokaryotes appear to possess numerous multi-type substrate-binding residues and are able to hierarchically activate these residues in a temperature-dependent manner, and thus act as temperature-regulated chaperones. The mechanism of hierarchical activation of substrate-binding residues is also discussed regarding its implication for eukaryotic sHSPs.

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IbpB‐bound substrate release in living cells as revealed by unnatural amino acid‐mediated photo‐crosslinking

Shi

Ezemaduka

2020

FEBS Open Bio

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Small heat shock proteins (sHSPs) are known to bind partially folded proteins under stress conditions. In this study, by using photo‐crosslinking, we showed that the substrate binding of sHSP IbpB in living cells is reversible upon short‐time exposure at elevated temperature, providing in vivo evidence on the existence of substrate protein release at nonstress conditions.

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In Vivo Substrate Diversity and Preference of Small Heat Shock Protein IbpB as Revealed by Using a Genetically Incorporated Photo-cross-linker

Cited by 54 publications

References 65 publications

A Small Heat Shock Protein Enables Escherichia coli To Grow at a Lethal Temperature of 50 C Conceivably by Maintaining Cell Envelope Integrity

A Small Heat Shock Protein Enables Escherichia coli To Grow at a Lethal Temperature of 50 C Conceivably by Maintaining Cell Envelope Integrity

Chaperone function and mechanism of small heat-shock proteins

IbpB‐bound substrate release in living cells as revealed by unnatural amino acid‐mediated photo‐crosslinking

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