Overexpression of (His)6-tagged human arginase I in Saccharomyces cerevisiae and enzyme purification using metal affinity chromatography

• Published in: Protein expression and purification Vol. 81; no. 1; pp. 63 - 68
• Main Authors: Zakalskiy, Andriy E., Zakalska, Oksana M., Rzhepetskyy, Yuriy A., Potocka, Natalia, Stasyk, Oleh V., Horak, Daniel, Gonchar, Mykhailo V.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.01.2012
• Subjects: (His)6-tag; Arginase - chemistry; Arginase - genetics; Arginase - isolation & purification; Arginase - metabolism; Chromatography, Affinity - methods; Cloning, Molecular; Histidine - chemistry; Histidine - genetics; Histidine - metabolism; Human arginase I; Humans; Liver - enzymology; Nitrilotriacetic Acid - analogs & derivatives; Oligopeptides - chemistry; Oligopeptides - genetics; Oligopeptides - metabolism; Organometallic Compounds; Protein purification; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - isolation & purification; Recombinant Fusion Proteins - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Human arginase I; (His)6-tag; Protein purification; Saccharomyces cerevisiae
• ISSN: 1046-5928; 1096-0279
• DOI: 10.1016/j.pep.2011.09.001

► (His)6-tagged human arginase I was expressed in the yeast Saccharomyces cerevisiae. ► The recombinant arginase was purified in one step by Nickel-affinity chromatography. ► The purified enzyme exhibits a specific activity up to 1600μmolmin−1mg−1 protein. Arginase (EC 3.5.3.1; l-arginine amidinohydrolase) is a key enzyme of the urea cycle that catalyses the conversion of arginine to ornithine and urea, which is the final cytosolic reaction of urea formation in the mammalian liver. The recombinant strain of the yeast Saccharomyces cerevisiae that is capable of overproducing arginase I (rhARG1) from human liver under the control of the efficient copper-inducible promoter CUP1, was constructed. The (His)6-tagged rhARG1 was purified in one step from the cell-free extract of the recombinant strain by metal-affinity chromatography with Ni–NTA agarose. The maximal specific activity of the 40-fold purified enzyme was 1600μmolmin−1mg−1 protein.