The hsp70 chaperone DnaK is a secondary amide peptide bond cis-trans isomerase.
Peptidyl prolyl cis-trans isomerases can enzymatically assist protein folding, but these enzymes exclusively target the peptide bond preceding proline residues. Here we report the identification of the Hsp70 chaperone DnaK as the first member of a novel enzyme class of secondary amide peptide bond cis-trans isomerases (APIases). APIases selectively accelerate the cis-trans isomerization of nonprolyl peptide bonds. Results from independent experiments support the APIase activity of DnaK: (i) exchange crosspeaks between the cis-trans conformers appear in 2D (1)H NMR exchange spectra of oligopeptides (ii) the rate constants for the cis-trans isomerization of various dipeptides increase and (iii) refolding of the RNase T1 P39A variant is catalyzed. The APIase activity shows both regio and stereo selectivity and is stimulated two-fold in the presence of the complete DnaK/GrpE/DnaJ/ATP refolding system. Moreover, known DnaK-binding oligopeptides simultaneously affect the APIase activity of DnaK and the refolding yield of denatured firefly luciferase in the presence of DnaK/GrpE/DnaJ/ATP. These results suggest a new role for the chaperone as a regioselective catalyst for bond rotation in polypeptides.
Schiene-Fischer
Cordelia
C
Max Planck Research Unit for Enzymology of Protein Folding, Weinbergweg 22, D-06120 Halle/Saale, Germany.
Habazettl
Judith
J
Schmid
Franz X
FX
Fischer
Gunter
G
D000255:Adenosine Triphosphate / Q000378:metabolism; D000577:Amides / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism; D001665:Binding Sites; D002384:Catalysis; D003080:Cold Temperature; D001517:Coleoptera; D004151:Dipeptides / Q000737:chemistry / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000378:metabolism; D007536:Isomerism; D007700:Kinetics; D008156:Luciferases / Q000737:chemistry / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008280:Magnetics; D017510:Protein Folding; D020673:Protein Renaturation; D006163:Ribonuclease T1 / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D019745:cis-trans-Isomerases / Q000378:metabolism*
D000577:Amides; D001426:Bacterial Proteins; D004151:Dipeptides; C494635:DnaJ protein, E coli; D029968:Escherichia coli Proteins; C068113:GrpE protein, Bacteria; C489400:GrpE protein, E coli; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D000255:Adenosine Triphosphate; D008156:Luciferases; D006163:Ribonuclease T1; C063842:dnaK protein, E coli; D019745:cis-trans-Isomerases