Purification of dopa decarboxylase from bovine striatum.
Pyridoxal phosphate-dependent DOPA decarboxylase has been purified from bovine striatum to a specific activity of 1.6 U/mg protein. After ammonium sulfate precipitation (30-60%) it was purified by DEAE-Sephacel, Sephacryl S-200, and TSK Phenyl 5 PW chromatography. The purified enzyme showed a single silver straining band with polyacrylamide gel electrophoresis under both denaturing and non-denaturing conditions. The bovine striatal DOPA decarboxylase is a dimer (subunit Mr = 56,000 by SDS-PAGE) with a native Mr of 106,000 as judged by chromatography on Sephacryl S-200 and by sedimentation analysis. Similar to the DOPA decarboxylase purified from non-CNS tissues, the bovine striatal enzyme requires free sulfhydryl groups for activity, is strongly inhibited by heavy metal ions, and can decarboxylate 5-hydroxytryptophan as well. It should be noted, however, that the final enzyme preparation is enriched in DOPA decarboxylase activity. The distribution of the DOPA decarboxylase and 5-HTP decarboxylase activities also varies among several bovine brain regions. In addition, heat treatment of the enzyme preparation inactivated the two decarboxylation activities at different rates.
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Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Manitoba, Winnipeg, Canada.
Dakshinamurti
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D006916:5-Hydroxytryptophan / Q000378:metabolism; D000645:Ammonium Sulfate; D000818:Animals; D001142:Aromatic-L-Amino-Acid Decarboxylases / Q000302:isolation & purification*; D002417:Cattle; D003342:Corpus Striatum / Q000201:enzymology*; D004295:Dihydroxyphenylalanine / Q000378:metabolism; D004296:Dopa Decarboxylase / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D008970:Molecular Weight; D011188:Potassium / Q000494:pharmacology; D012995:Solubility; D013438:Sulfhydryl Compounds / Q000494:pharmacology
D013438:Sulfhydryl Compounds; D004295:Dihydroxyphenylalanine; D006916:5-Hydroxytryptophan; D004296:Dopa Decarboxylase; D001142:Aromatic-L-Amino-Acid Decarboxylases; D011188:Potassium; D000645:Ammonium Sulfate