A functional DnaK dimer is essential for the efficient interaction with Hsp40 heat shock protein
| Autor: | Jiao Yang, Qinglian Liu, Evans Boateng Sarbeng, Jennifer Li Wong, Hongtao Li, Qingdai Liu, Lei Zhou, Xueli Tian |
|---|---|
| Rok vydání: | 2014 |
| Předmět: |
Dimer
genetic processes Allosteric regulation Fluorescence Polarization Plasma protein binding Biochemistry Oligomer Protein–protein interaction chemistry.chemical_compound Heat shock protein HSP70 Heat-Shock Proteins Molecular Biology Escherichia coli Proteins Cell Biology HSP40 Heat-Shock Proteins Surface Plasmon Resonance Proteostasis chemistry biological sciences Protein Structure and Folding Mutagenesis Site-Directed bacteria Protein folding Dimerization Protein Binding |
| Zdroj: | The Journal of biological chemistry. 290(14) |
| ISSN: | 1083-351X |
| Popis: | Highly conserved molecular chaperone Hsp70 heat shock proteins play a key role in maintaining protein homeostasis (proteostasis). DnaK, a major Hsp70 in Escherichia coli, has been widely used as a paradigm for studying Hsp70s. In the absence of ATP, purified DnaK forms low-ordered oligomer, whereas ATP binding shifts the equilibrium toward the monomer. Recently, we solved the crystal structure of DnaK in complex with ATP. There are two molecules of DnaK-ATP in the asymmetric unit. Interestingly, the interfaces between the two molecules of DnaK are large with good surface complementarity, suggesting functional importance of this crystallographic dimer. Biochemical analyses of DnaK protein supported the formation of dimer in solution. Furthermore, our cross-linking experiment based on the DnaK-ATP structure confirmed that DnaK forms specific dimer in an ATP-dependent manner. To understand the physiological function of the dimer, we mutated five residues on the dimer interface. Four mutations, R56A, T301A, N537A, and D540A, resulted in loss of chaperone activity and compromised the formation of dimer, indicating the functional importance of the dimer. Surprisingly, neither the intrinsic biochemical activities, the ATP-induced allosteric coupling, nor GrpE co-chaperone interaction is affected appreciably in all of the mutations except for R56A. Unexpectedly, the interaction with co-chaperone Hsp40 is significantly compromised. In summary, this study suggests that DnaK forms a transient dimer upon ATP binding, and this dimer is essential for the efficient interaction of DnaK with Hsp40. |
| Databáze: | OpenAIRE |
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