New approaches to chromatographic purification of bovine dopamine-beta-hydroxylase.
The use of the traditional scheme for the isolation of bovine dopamine-beta-hydroxylase (bDBH) from bovine adrenal medulla resulted in active but not pure bDBH, containing about 50% of admixtures. Immobilized metal chelate affinity chromatography on agarose modified with iminodiacetic acid residues and charged with cobalt ions was applied in the final stage to obtain more than 90% pure and active bDBH. Final purification of bDBH using step elution with 0-0.5 M methyl-D-mannoside in buffer solution from concanavalin A-Sepharose was studied. The determination of bDBH in various samples was performed using size-exclusion chromatography.
D000313:Adrenal Medulla / Q000201:enzymology; D000818:Animals; D002417:Cattle; D002845:Chromatography / Q000379:methods*; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004299:Dopamine beta-Hydroxylase / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D013056:Spectrophotometry, Ultraviolet
D004299:Dopamine beta-Hydroxylase