Purification and characterization of chemotactic methylesterase from Bacillus subtilis

Abstract

By utilization of methanol evolution as an assay, a protein methylesterase from Bacillus subtilis has been purified. A 1200-fold purification has been achieved by CM-Bio-Gel A, hydroxylapatite, and Bio-Gel P-60 column chromatography. Gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicate the enzyme is a monomer of 41 000 in molecular weight. The enzyme is stabilized and activated by aqueous glycerol solutions. Methyl-accepting chemotaxis proteins (MCPs) serve as substrates for the enzyme. The enzyme requires divalent cation for activity, with maximum activity obtained at 1.1 mM Mg2+. The enzyme is most active at pH 7.5 and at 28 degrees C. Methylesterase has an apparent Km for methylated MCPs of about 10 nM.