Soluble adenylate cyclase from the culture medium of Bordetella pertussis: purification and characterization

Abstract

Culture medium of exponentially growing B. pertussis (strain 114) contains significant quantities of soluble (100,000 .times. g for 1 h) adenylate cyclase. The enzyme was purified by chromatography on DEAE cellulose and Sephadex G-200. The purest material yielded a single band on sodium dodecyl sulfate-disc gel electrophoresis. It is heat labile, has a temperature optimum of 30.degree. C, a pH optimum of pH 7-8, a Km for ATP of 0.4 mM and requires Mg2+ for maximum activity. The MW, by sodium dodecyl sulfate-disc gel electrophoresis and sucrose density gradient, is .apprx. 70,000. The enzyme is markedly inhibited by fluoride and weakly inhibited by monovalent salts, but its activity is not altered by .alpha.-keto acids or nonsubstrate nucleoside triphosphates. This enzyme differs from the bacterial adenylate cyclases previously described.