Thermodynamic Linkage in the GrpE Nucleotide Exchange Factor, a Molecular Thermosensor

Gelinas, Amy D.; Toth, Joseph; Bethoney, Kelley A.; Langsetmo, Knut; Stafford, Walter F.; Harrison, Celia J.

doi:10.1021/bi034416b

Cited by 27 publications

(31 citation statements)

References 16 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…A similar deletion variant was able to refold thermally denatured luciferase in the DnaK system as efficiently as wtGrpE (34), indicating that the chaperone activity depends on the conformational properties and stability of the unfolded and aggregated substrates. When GrpE(69 -197) was used as NEF, the initial refolding rate was reduced by 65% (1.6 Ϯ 0.2% min Ϫ1 ), coinciding with data reported for the shorter GrpE(89 -197) construct (38). Based on the ability of the GrpE(69 -197) mutant to dissociate DnaK-substrate complexes (7), premature release of unfolded luciferase might hamper correct refolding of the substrate.…”

Section: Dnak-grpe Complex Formation With the Dnak And Grpesupporting

confidence: 80%

Modulation of the Chaperone DnaK Allosterism by the Nucleotide Exchange Factor GrpE

Melero

Moro

Pérez-Calvo

et al. 2015

Journal of Biological Chemistry

View full text Add to dashboard Cite

Background: DnaK is a Hsp70 (heat shock protein) molecular chaperone whose function is controlled in part by the nucleotide exchange factor GrpE. Results: We describe the structures of several complexes formed between GrpE and different DnaK variants. Conclusion: The DnaK-GrpE complex has several conformations, an important factor in regulating DnaK function. Significance: The information obtained explains the nucleotide exchange role of GrpE in much more detail.

show abstract

Section: Dnak-grpe Complex Formation With the Dnak And Grpesupporting

confidence: 80%

Modulation of the Chaperone DnaK Allosterism by the Nucleotide Exchange Factor GrpE

Melero

Moro

Pérez-Calvo

et al. 2015

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The first part of the temperature-induced structural transition of GrpE Eco has been attributed to the unfolding of the N-terminal long paired helices, 13 which is linked to the destabilization of the β-sheet domain. 14 In the case of GrpE Tth , the lower-temperature transition with a midpoint at 90°C has been alternatively ascribed to the unfolding of the Cterminal β-sheet domain, not the N-terminal helices. 8 Meanwhile, the second transition (at 75-80°C in GrpE Eco 7 and at 100-105°C in GrpE Tth 8 ) has been considered to be responsible for the irreversible dissociation of the GrpE dimer in both species.…”

Section: Introductionmentioning

confidence: 99%

Crystal Structure of a Thermophilic GrpE Protein: Insight into Thermosensing Function for the DnaK Chaperone System

Nakamura

Takumi

Miki

2010

Journal of Molecular Biology

View full text Add to dashboard Cite

A homodimeric GrpE protein functions as a nucleotide exchange factor of the eubacterium DnaK molecular chaperone system. The co-chaperone GrpE accelerates ADP dissociation from, and promotes ATP binding to, DnaK, which cooperatively facilitates the DnaK chaperone cycle with another co-chaperone, DnaJ. GrpE characteristically undergoes two-step conformational changes in response to elevation of the environmental temperature. In the first transition at heat-shock temperatures, a fully reversible and functionally deficient structural alteration takes place in GrpE, and then the higher temperatures lead to the irreversible dissociation of the GrpE dimer into monomers as the second transition. GrpE is also thought to be a thermosensor of the DnaK system, since it is the only member of the DnaK system that changes its structure reversibly and loses its function at heat-shock temperatures of various organisms. We here report the crystal structure of GrpE from Thermus thermophilus HB8 (GrpE(Tth)) at 3.23 A resolution. The resolved structure is compared with that of GrpE from mesophilic Escherichia coli (GrpE(Eco)), revealing structural similarities, particularly in the DnaK interaction regions, and structural characteristics for the thermal stability of GrpE(Tth). In addition, the structure analysis raised the possibility that the polypeptide chain in the reported GrpE(Eco) structure was misinterpreted. Comparison of these two GrpE structures combined with the results of limited proteolysis experiments provides insight into the protein dynamics of GrpE(Tth) correlated with the shift of temperature, and also suggests that the localized and partial unfolding at the plausible DnaK interaction sites of GrpE(Tth) causes functional deficiency of nucleotide exchange factor in response to the heat shock.

show abstract

“…Stabilization of the pair of the long NH 2 -terminal helices in the GrpE dimer with an engineered disulfide bond (R40C) (Fig. 2) abolishes the thermal transition in GrpE and reduces the deviation of the ADP/ATP exchange activity from an Arrhenius temperature dependence, indicating that the long helix pair acts as the primary thermosensor of the chaperone system (15,16).In E. coli, DnaK and its co-chaperones are constitutively expressed as well as induced by heat shock or other cellular stress. This familiar heat shock response is mediated by 32 , which directs RNA polymerase to transcribe the particular set of heat shock genes.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

The Importance of Having Thermosensor Control in the DnaK Chaperone System

Siegenthaler

Christen

2005

Journal of Biological Chemistry

View full text Add to dashboard Cite

In addition to the 32 -mediated heat shock response, the DnaK/DnaJ/GrpE molecular chaperone system of Escherichia coli directly adapts to elevated temperatures by sequestering a higher fraction of substrate. This immediate heat shock response is due to the differential temperature dependence of the activity of DnaJ, which stimulates the hydrolysis of DnaK-bound ATP, and the activity of GrpE, which facilitates ADP/ATP exchange and converts DnaK from its high-affinity ADPliganded state into its low-affinity ATP-liganded state. GrpE acts as thermosensor with its ADP/ATP exchange activity decreasing above 40°C. To assess the importance of this reversible thermal adaptation for the chaperone action of the DnaK/DnaJ/GrpE system during heat shock, we used glucose-6-phosphate dehydrogenase and luciferase as substrates. We compared the performance of wild-type GrpE as a component of the chaperone system with that of GrpE R40C. In this mutant, the thermosensing helices are stabilized with an intersubunit disulfide bond and its nucleotide exchange activity thus increases continuously with increasing temperature. Wild-type GrpE with intact thermosensor proved superior to GrpE R40C with desensitized thermosensor. The chaperone system with wild-type GrpE yielded not only a higher fraction of refolding-competent protein at the end of a heat shock but also protected luciferase more efficiently against inactivation during heat shock. Consistent with their differential thermal behavior, the protective effects of wild-type GrpE and GrpE R40C diverged more and more with increasing temperature. Thus, the direct thermal adaptation of the DnaK chaperone system by thermosensing GrpE is essential for efficient chaperone action during heat shock.Molecular chaperones of the 70-kDa heat shock protein (Hsp70) 1 family participate in many cellular processes, including the folding, membrane translocation, and degradation of proteins (1). The chaperones recognize and interact with hydrophobic peptide segments, which are exposed by nascent polypeptide chains during synthesis and by misfolded proteins during stress, in particular heat shock. The irreversible formation of protein aggregates thus is reduced, increasing the yield of properly folded and refolded native protein. Hsp70 bind and release their substrates in an ATP-driven cycle (2, 3). The ATPase activity resides in the NH 2 -terminal domain (4, 5) and modulates the substrate binding properties of the COOH-terminal peptide-binding domain (6). The ATP-liganded T state is characterized by low affinity for substrates and fast rates of binding and release, whereas the ADP-liganded R state shows high affinity for substrates with slow kinetics (7,8). DnaK, an Hsp70 homolog in Escherichia coli, acts in concert with its co-chaperones DnaJ, an Hsp40 homolog, and GrpE (2, 9, 10). DnaJ stimulates the hydrolysis of DnaK-bound ATP and converts T-state DnaK into its high-affinity R state (Fig. 1). GrpE facilitates ADP/ATP exchange and reconverts DnaK into the low-affinity T state. In the presence of A...

show abstract

Thermodynamic Linkage in the GrpE Nucleotide Exchange Factor, a Molecular Thermosensor

Cited by 27 publications

References 16 publications

Modulation of the Chaperone DnaK Allosterism by the Nucleotide Exchange Factor GrpE

Modulation of the Chaperone DnaK Allosterism by the Nucleotide Exchange Factor GrpE

Crystal Structure of a Thermophilic GrpE Protein: Insight into Thermosensing Function for the DnaK Chaperone System

The Importance of Having Thermosensor Control in the DnaK Chaperone System

Contact Info

Product

Resources

About