Abstract
A DNA-dependent RNA polymerase has been isolated from Caldariella acidophila, a thermophilic bacterium living in acidic hot springs at temperatures ranging from 63 to 89 degrees C. The enzyme was purified 180-fold and is composed of five different subunits having the following molecular weights: a = 127000, b = 120000, c = 72000, d = 65000, and e = 38000. The enzyme is activated by Mn2+ and Mg2+ and exhibits optimal activity in the presence of 0.5 mM Mn2+. The activity depends on ionic strength, with a maximum at 0.25 M KCl, and exhibits a pH optimum at 7.8 in the presence of Tris-HCl buffer. The enzyme shows a high degree of thermophilicity, its temperature optimum being 80 degrees C in the in vitro assay. The thermophilicity of C. acidophila RNA polymerase allows studies on enzyme-template interactions to be performed in a temperature range where many templates are close to their Tm.
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