A new enzyme, L-ribose isomerase, was found constitutively in Acinetobacter sp. strain DL-28. The enzyme was purified by polyethylene glycol precipitation, anion exchange chromatography on diethylaminoethyl (DEAE)-Toyopearl 650M and gel filtration on Sephadex G-150. The enzyme was found to be homogenous by polyacrylamide gel electrophoresis. The molecular weight of the enzyme was estimated to be about 120,000 by Sephadex G-150 gel filtration. On the basis of sodium dodecyl sulfate (SDS) gel electrophoresis, the enzyme is composed of four identical subunits with molecular weights of about 32,000. The isoelectric point of the enzyme was estimated to be about 5.1. The enzyme was specific for L-ribose, D-lyxose and D-mannose. The K-m for L-ribose was 44 mM and Y-max was 357 mu mol/mg . min. The equilibrium ratio between L-ribose and L-ribulose was 70:30. The maximum activity at 30 degrees C was obtained at pH 9.0, and the enzyme was stable in the pH range of 7.0-9.0. The optimum temperature was around 30 degrees C, and the enzyme was stable up to 30 degrees C for 10 min.