Probing the transient interaction between the small heat-shock protein Hsp21 and a model substrate protein using crosslinking mass spectrometry

Lambert, Wietske; Rutsdottir, Gudrun; Hussein, Rasha M.; Bernfur, Katja; Kjellström, Sven; Emanuelsson, Cecilia

doi:10.1007/s12192-012-0360-4

Cited by 15 publications

(23 citation statements)

References 47 publications

(56 reference statements)

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“…However, the truncations also significantly affect the structural integrity of sHSPs and therefore, the exact roles of the corresponding regions for substrate-binding could not be unambiguously assigned. A more reliable strategy based on chemical crosslinking coupled with mass spectrometry was successfully applied to the characterization of the substrate-binding regions of M. tuberculosis Hsp16.3, plant Hsp21, and mammalian aA-crystallin, aB-crystallin, and Hsp22 [105][106][107][108][109], revealing that the N-terminal part of the a-crystallin domain and the N-terminal arm are the major regions for substrate binding. Notably, these substrate-binding regions appear to contain both polar and non-polar amino acids.…”

Section: Multi-type Residues Of Shsps Participate In Binding Substratmentioning

confidence: 99%

Chaperone function and mechanism of small heat-shock proteins

Fu¹

2014

ABBS

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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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Section: Multi-type Residues Of Shsps Participate In Binding Substratmentioning

confidence: 99%

Chaperone function and mechanism of small heat-shock proteins

Fu¹

2014

ABBS

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“…Whereas the exact role of the fl exible C-terminal extension in binding substrate proteins is still in debate (Carver and Lindner 1998 ;Smulders et al 1996 ;Jehle et al 2010 ;Saji et al 2008 ;Treweek et al 2007 ;Jaya et al 2009 ;Jiao et al 2005b ;Fu et al 2013b ), it is well established that the structurally disordered N-terminal arm serves as a principal substrate-binding region (Fu et al , 2013bSharma et al 1997 ;Jaya et al 2009 ;Lambert et al 2013 ). In this context, sHSPs appear to dominantly utilize their disordered N-terminal arms for substrate-binding, and thus allow them to present diverse geometries for substrate-binding.…”

Section: Disorder-adapting-disorder For Shsp-substrate Recognition Anmentioning

confidence: 99%

“…Studies by truncation or by chemical crosslinking showed that the three characteristic domains of sHSPs are all crucial for the chaperone functional integrity (Merck et al 1993b ;Fu and Chang 2006a ;Fu et al 2005 ;Basha et al 2006 ;Giese and Vierling 2004 ;Leroux et al 1997b ;Jiao et al 2005b ;Lentze et al 2003 ;Ghosh et al 2006 ;Saji et al 2008 ;Strozecka et al 2012 ;Studer et al 2002 ;Treweek et al 2007 ;Fu and Chang 2006b ;Lambert et al 2013 ;Shemetov and Gusev 2011 ). However, these studies cannot reveal the exact role of each amino acid residue for substrate-binding.…”

Section: Structural Diversity and Spatial Chaos Of Substratebinding Rmentioning

confidence: 99%

Insights into How Small Heat Shock Proteins Bind a Great Diversity of Substrate Proteins: A Super-Transformer Model

2015

Heat Shock Proteins

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Small heat shock proteins (sHSPs) are ubiquitous, ATP-independent molecular chaperones that play crucial roles in the cellular protein quality control and whose dysfunction is also linked to various diseases. They are able to suppress the aggregation of non-native substrate proteins and assist the refolding of these substrates in cooperation with ATP-dependent chaperones. Substrate recognition and binding by sHSPs are essential for their chaperone functions. In this review, I focus on how a sHSP recognizes and binds various substrate proteins that are greatly diversifi ed in both structures and functions. Such a broad substrate spectrum is bestowed not only by the well-known dynamics of sHSP oligomeric structures, but also by their characteristic disordered substrate-binding regions that may interact with disordered substrate proteins via a way of disorder-adapting-disorder. Further, utilization of numerous substrate-binding residues is also crucial, as these residues include both hydrophobic and polar amino acids equally and are also spatially chaotic throughout the sHSP oligomers. In keeping with these details, a novel super-transformer model is presented to account for the structural characteristics and broad substrate spectrum of sHSPs.

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“…We have repeatedly seen in proteolysis experiments that the N-terminal domain in Hsp21 is rapidly degraded, behaving as intrinsically disordered proteins (Bardwell and Jakob 2012 ;Tompa and Csermely 2004 ) and in various functional in vitro assays Hsp21 displays the typical properties of a molecular chaperone, suppressing the aggregation of heat-sensitive model substrate proteins and of fi brillation-prone peptides (unpublished data). Chemical crosslinking and mass spectrometry suggest an interaction between the disordered N-terminal region of Hsp21 and the C-terminal presumably unfolding part of the temperature-sensitive substrate protein malate dehydrogenase (Lambert et al 2013 ).…”

Section: The Chloroplast-localized Shsp Hsp21mentioning

confidence: 99%

The Chloroplast-Localized Plant sHsp in Arabidopsis Thaliana: Role of Its Oligomeric Conformation and Its Translocation into Membranes

Bernfur

Rutsdottir

Månsson

et al. 2015

Heat Shock Proteins

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Recent advances in quantitative proteomics show that small heat shock proteins (sHsps) are among the most highly upregulated proteins in cellular stress response. In plants there are numerous paralogous sHsps expressed in various cellular compartments. The chloroplast-localized sHsp, named Hsp21 with reference to its monomeric size, has an N-terminal region that is some 40 amino acids longer compared to the cytosolic sHsps, and increases plant stress tolerance as shown by Arabidopsis thaliana plants which overexpress Hsp21. Recombinantly expressed and purifi ed Hsp21-protein shows the features expected for a chaperone protein in rescuing temperature-sensitive model substrate proteins from aggregation. Hsp21 is a dodecameric protein, with C-terminal tails that keep the dodecamer together but are highly fl exible in solution. The N-terminal domains, which resemble intrinsically disordered proteins and are located in the interior of the dodecamer, contain conserved methionine residues of crucial importance for function. Methionine sulfoxidation abolishes the chaperone activity in vitro, but in vivo such oxidized methioniones appear to be continuously re-reduced thanks to a chloroplast-localized form of peptide methionine sulfoxide reductase, an important enzyme expressed ubiquitously and belonging to the minimal gene set for life. There is not a clear picture of which the endogenous substrate proteins of Hsp21 are. For more than a decade it has been observed that cyanobacterial sHsps enter the membranes at increased temperatures. In a quantitative proteomics approach we have recently analyzed and compared Hsp21 with hundreds of other chloroplast proteins, and found that Hsp21 is fairly unique with respect to its translocation into membrane in heat-stressed plants. One reason for the oligomericity of Hsp21 is suggested to be the possibility to rapidly supply hydrophobic surfaces with chaperoning capacity in response to stress, while still preventing membrane lysis by such hydrophobic surfaces under non-stress conditions.

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Probing the transient interaction between the small heat-shock protein Hsp21 and a model substrate protein using crosslinking mass spectrometry

Cited by 15 publications

References 47 publications

Chaperone function and mechanism of small heat-shock proteins

Chaperone function and mechanism of small heat-shock proteins

Insights into How Small Heat Shock Proteins Bind a Great Diversity of Substrate Proteins: A Super-Transformer Model

The Chloroplast-Localized Plant sHsp in Arabidopsis Thaliana: Role of Its Oligomeric Conformation and Its Translocation into Membranes

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