Role of DnaJ G/F-rich domain in conformational recognition and binding of protein substrates.
DnaJ from Escherichia coli is a Type I Hsp40 that functions as a cochaperone of DnaK (Hsp70), stimulating its ATPase activity and delivering protein substrates. How DnaJ binds protein substrates is still poorly understood. Here we have studied the role of DnaJ G/F-rich domain in binding of several substrates with different conformational properties (folded, partially (un)folded and unfolded). Using partial proteolysis we find that RepE, a folded substrate, contacts a wide DnaJ area that involves part of the G/F-rich region and Zn-binding domain. Deletion of G/F-rich region hampers binding of native RepE and reduced the affinity for partially (un)folded substrates. However, binding of completely unfolded substrates is independent on the G/F-rich region. These data indicate that DnaJ distinguishes the substrate conformation and is able to adapt the use of the G/F-rich region to form stable substrate complexes.
Perales-Calvo
Judit
J
Unidad de Biofísica (CSIC/UPV-EHU), Departamento de Bioquímica y Biología Molecular, Facultad de Ciencia y Tecnología, Universidad del País Vasco, Apartado 644, 48080 Bilbao, Spain.
Muga
Arturo
A
Moro
Fernando
F
D016880:Anisotropy; D018833:Chaperonins / Q000737:chemistry; D003001:Cloning, Molecular; D003432:Cross-Linking Reagents / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000737:chemistry*; D050956:HSP40 Heat-Shock Proteins / Q000737:chemistry*; D057927:Hydrophobic and Hydrophilic Interactions; D013058:Mass Spectrometry / Q000379:methods; D009154:Mutation; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D014357:Trypsin / Q000737:chemistry
D003432:Cross-Linking Reagents; C494635:DnaJ protein, E coli; D029968:Escherichia coli Proteins; D050956:HSP40 Heat-Shock Proteins; D014357:Trypsin; D018833:Chaperonins