Crystalline L-ribulokinase from Escherichia coli.

r.-Ribulokinase has been purified and crystallized from induced cultures of Escherichia coli B/r mutant araA-2, a hyperproducer of this enzyme. The enzyme appears homogeneous by Sephadex G-200 column chromatography and sedimentation pattern. Recrystallization does not increase its specific activity. Acrylamide gel disc electrophoresis at pH 8.3 shows a trace of a slower component, not exceeding 5 % of the total material. The molecular weight of L-ribulokinase as determined by osmotic pressure, sedimentation equilibrium, and sedimentation and diffusion is 9.8 f 0.2 X 104. The enzyme is unstable at acid pH values, but is stable when stored at pH 7.6 or above. It resists inactivation by heating at 60.5” for 30 min at this pH. It phosphorylates both Land D-ribulose, the K, for n-ribulose being about twice that for L-ribulose. The molecular activity for purified enzyme is about 1700 moles of product formed per min per mole of enzyme for L-ribulose. Amino acid analysis shows the absence of valine and methionine. The enzyme is estimated to occupy about 3 to 4% of the total protein in the crude extract of the fully induced araA-2 cell.