An improvement in the pyruvate dehydrogenase complex assay: a high-yield method for purifying arylamine acetyltransferase.
A new method for purifying arylamine acetyltransferase (AAT) has been devised using polyethylene glycol fractionation and hydroxylapatite chromatography. The new procedure gives a final yield of approximately 70% based on activity in crude homogenates and can be performed in a single day. This represents a threefold higher yield than previous methods. The procedure may be used to purify AAT from pigeon, chicken, and duck livers with equivalent yield of the final enzyme. However, the preparation from pigeon liver is preferred because of the sevenfold higher activity in this tissue. Nevertheless, if fresh pigeon livers cannot be acquired, calculations reveal that the preparation from chicken liver is comparable to that from pigeon liver acetone powder with respect to total activity obtained per gram wet weight starting material. One can also calculate that the chicken liver preparation is approximately 40 times cheaper than that from pigeon liver acetone powder, making the preparation of AAT from fresh chicken livers a good alternative when pigeon livers are not available.
Brooks
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SP
Nutrition Research Division, Health and Welfare Canada, Banting Research Centre, Tunney's Pasture, Ottawa, Ontario.
Storey
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KB
D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D001191:Arylamine N-Acetyltransferase / Q000302:isolation & purification*; D001391:Azo Compounds / Q000378:metabolism; D002645:Chickens; D010856:Columbidae; D004372:Ducks; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000201:enzymology; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism*; D012967:Sodium Dodecyl Sulfate; D013425:Sulfanilic Acids / Q000378:metabolism
D000814:Aniline Compounds; D001391:Azo Compounds; D011768:Pyruvate Dehydrogenase Complex; D013425:Sulfanilic Acids; C083612:4-(4-aminophenylazo)benzenesulfonic acid; C019498:4-nitroaniline; D012967:Sodium Dodecyl Sulfate; D001191:Arylamine N-Acetyltransferase