FIG. 4.

Kinetics of peptide release by DnaK and DnaK derivatives. DnaK or mutant DnaK protein (1 μM) was incubated with PepH-ANBD (0.5 μM) in ATPase buffer for 30 min at 30°C and mixed in a stopped-flow apparatus with an equal volume of ATP (2 μM) in ATPase buffer. The excitation wavelength used was 480 nm, and a 495-nm-cutoff filter was used. For details of fluorescence measurements, see Materials and Methods. All reaction traces, except for that of DnaK K70A, could be fit to a single-exponential decay function.