FIG. 6.

The presence of a substrate peptide inhibits autophosphorylation. The DE-52 fraction, 20 μg, was incubated for 60 min at 65°C in a volume of 100 μl of 20 mM MES, pH 6.5, containing 50 μM ATP (300 μCi of [γ-³²P]ATP/ml), 5 mM MnCl₂, 2 mM DTT, 12.5 mM octyl glucoside, and the indicated concentrations of the MLC peptide. The reaction was stopped by precipitating the proteins with 3 volumes of cold acetone. The precipitated proteins were collected by centrifugation and resuspended in SDS-sample buffer and analyzed by SDS-PAGE using an 8% (wt/vol) gel. Shown is an autoradiogram of the gel. Arrow, position of the S. solfataricus protein kinase. The concentration of the MLC peptide present in each sample (millimolar) is indicated at the top of each lane.