Control of the DnaK chaperone cycle by substoichiometric concentrations of the co-chaperones DnaJ and GrpE
The polypeptide binding and release cycle of the molecular chaperone DnaK (Hsp70) of Escherichia coli is regulated by the two co-chaperones DnaJ and GrpE. Here, we show that the DnaJ-triggered conversion of DnaK.ATP (T state) to DnaK.ADP.Pi (R state), as monitored by intrinsic protein fluorescence, is monophasic and occurs simultaneously with ATP hydrolysis. This is in contrast with the T-->R conversion in the absence of DnaJ which is biphasic, the first phase occurring simultaneously with the hydrolysis of ATP (Theyssen, H., Schuster, H.-P., Packschies, L., Bukau, B., and Reinstein, J. (1996) J. Mol. Biol. 263, 657-670). Apparently, DnaJ not only stimulates ATP hydrolysis but also couples it with conformational changes of DnaK. In the absence of GrpE, DnaJ forms a tight ternary complex with peptide.DnaK.ADP.Pi (Kd = 0.14 microM). However, by monitoring complex formation between DnaK (1 microM) and a fluorophore-labeled peptide in the presence of ATP (1 mM), DnaJ (1 microM), and varying concentrations of the ADP/ATP exchange factor GrpE (0.1-3 microM), substoichiometric concentrations of GrpE were found to shift the equilibrium from the slowly binding and releasing, high-affinity R state of DnaK completely to the fast binding and releasing, low-affinity T state and thus to prevent the formation of a long lived ternary DnaJ. substrate.DnaK.ADP.Pi complex. Under in vivo conditions with an estimated chaperone ratio of DnaK:DnaJ:GrpE = 10:1:3, both DnaJ and GrpE appear to control the chaperone cycle by transient interactions with DnaK.
| Medienart: |
Artikel |
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| Erscheinungsjahr: |
1998 |
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| Erschienen: |
1998 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:273 |
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| Enthalten in: |
The Journal of biological chemistry - 273(1998), 12 vom: 20. März, Seite 6643-9 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Pierpaoli, E V [VerfasserIn] |
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| Anmerkungen: |
Date Completed 16.04.1998 Date Revised 09.02.2021 published: Print Citation Status MEDLINE |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM094486778 |
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| 041 | |a eng | ||
| 100 | 1 | |a Pierpaoli, E V |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Control of the DnaK chaperone cycle by substoichiometric concentrations of the co-chaperones DnaJ and GrpE |
| 264 | 1 | |c 1998 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ohne Hilfsmittel zu benutzen |b n |2 rdamedia | ||
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| 500 | |a Date Completed 16.04.1998 | ||
| 500 | |a Date Revised 09.02.2021 | ||
| 500 | |a published: Print | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a The polypeptide binding and release cycle of the molecular chaperone DnaK (Hsp70) of Escherichia coli is regulated by the two co-chaperones DnaJ and GrpE. Here, we show that the DnaJ-triggered conversion of DnaK.ATP (T state) to DnaK.ADP.Pi (R state), as monitored by intrinsic protein fluorescence, is monophasic and occurs simultaneously with ATP hydrolysis. This is in contrast with the T-->R conversion in the absence of DnaJ which is biphasic, the first phase occurring simultaneously with the hydrolysis of ATP (Theyssen, H., Schuster, H.-P., Packschies, L., Bukau, B., and Reinstein, J. (1996) J. Mol. Biol. 263, 657-670). Apparently, DnaJ not only stimulates ATP hydrolysis but also couples it with conformational changes of DnaK. In the absence of GrpE, DnaJ forms a tight ternary complex with peptide.DnaK.ADP.Pi (Kd = 0.14 microM). However, by monitoring complex formation between DnaK (1 microM) and a fluorophore-labeled peptide in the presence of ATP (1 mM), DnaJ (1 microM), and varying concentrations of the ADP/ATP exchange factor GrpE (0.1-3 microM), substoichiometric concentrations of GrpE were found to shift the equilibrium from the slowly binding and releasing, high-affinity R state of DnaK completely to the fast binding and releasing, low-affinity T state and thus to prevent the formation of a long lived ternary DnaJ. substrate.DnaK.ADP.Pi complex. Under in vivo conditions with an estimated chaperone ratio of DnaK:DnaJ:GrpE = 10:1:3, both DnaJ and GrpE appear to control the chaperone cycle by transient interactions with DnaK | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 7 | |a Bacterial Proteins |2 NLM | |
| 650 | 7 | |a DnaJ protein, E coli |2 NLM | |
| 650 | 7 | |a Escherichia coli Proteins |2 NLM | |
| 650 | 7 | |a GrpE protein, Bacteria |2 NLM | |
| 650 | 7 | |a GrpE protein, E coli |2 NLM | |
| 650 | 7 | |a HSP40 Heat-Shock Proteins |2 NLM | |
| 650 | 7 | |a HSP70 Heat-Shock Proteins |2 NLM | |
| 650 | 7 | |a Heat-Shock Proteins |2 NLM | |
| 650 | 7 | |a Molecular Chaperones |2 NLM | |
| 650 | 7 | |a Adenosine Triphosphate |2 NLM | |
| 650 | 7 | |a 8L70Q75FXE |2 NLM | |
| 650 | 7 | |a dnaK protein, E coli |2 NLM | |
| 650 | 7 | |a EC 3.6.1.- |2 NLM | |
| 700 | 1 | |a Sandmeier, E |e verfasserin |4 aut | |
| 700 | 1 | |a Schönfeld, H J |e verfasserin |4 aut | |
| 700 | 1 | |a Christen, P |e verfasserin |4 aut | |
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| 773 | 1 | 8 | |g volume:273 |g year:1998 |g number:12 |g day:20 |g month:03 |g pages:6643-9 |
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| 952 | |d 273 |j 1998 |e 12 |b 20 |c 03 |h 6643-9 | ||