Bordetella pertussis adenylate cyclase: isolation and purification by calmodulin-sepharose 4B chromatography.
Purified preparations of adenylate cyclase were obtained from crude urea extracts of Bordetella pertussis by a one-step calmodulin affinity chromatography technique. Diluted extract was loaded onto the column and washed, and adenylate cyclase was eluted with 10mM EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid]. A 104-fold purification was accomplished in one step. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the affinity-purified adenylate cyclase was dissociated into one major protein band with an apparent molecular weight of 60,000 and a minor band at 200,000. The affinity-purified adenylate cyclase was observed to have adenylate cyclase enzymatic activity which was activated by calmodulin, to bind 125I-calmodulin, and to be free of pertussis toxin as determined by in vivo and in vitro assays.
D037361:Adenylate Cyclase Toxin; D000262:Adenylyl Cyclases / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D001886:Bordetella pertussis / Q000201:enzymology* / Q000472:pathogenicity; D002147:Calmodulin / Q000378:metabolism; D002449:Cell Aggregation / Q000187:drug effects; D002846:Chromatography, Affinity / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D051379:Mice; D009504:Neutrophils / Q000187:drug effects; D037342:Pertussis Toxin; D010566:Virulence Factors, Bordetella / Q000032:analysis
D037361:Adenylate Cyclase Toxin; D002147:Calmodulin; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D000262:Adenylyl Cyclases