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Xanthine oxidase and aldehyde oxidase: a simple procedure for the simultaneous purification from rat liver.
Arch Biochem Biophys. 2002 Apr 01; 400(1):48-53.AB

Abstract

Aldehyde oxidase (AO) and xanthine oxidase (XO) are cytosolic enzymes that have been involved in some pathological conditions and play an important role in the biotransformation of drugs and xenobiotics. The increasing interest in these enzymes demands for a simple and rapid procedure for their purification. This paper describes for the first time a method that allows simultaneous purification of both enzymes from the same batch of rat livers. It involves few steps, is reproducible and offers high enzyme yields with high specific activities. The rat liver homogenate was fractionated by heat denaturation and by ammonium sulphate precipitation to give a crude extract containing both enzymes. This extract was chromatographed on an Hydroxyapatite column that completely separated AO from XO. Further purification of XO by anion exchange chromatography on a Q-Sepharose Fast Flow column resulted in a highly purified (1200-fold) preparation, with a specific activity of 3.64 U/mg and with a 20% yield. AO was purified about 1000-fold at a yield of 15%, with a specific activity of 3.48 U/mg, by affinity chromatography on Benzamidine-Sepharose 6B. The purified enzymes gave single bands of approximately 300 kDa on a polyacrylamide gel gradient electrophoresis and displayed the characteristic absorption spectra of highly purified enzymes.

Authors+Show Affiliations

Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade de Lisboa, Edifício C8, Campo Grande, 1749-016 Lisboa, Portugal.No affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

11913970

Citation

Maia, Luisa, and Lurdes Mira. "Xanthine Oxidase and Aldehyde Oxidase: a Simple Procedure for the Simultaneous Purification From Rat Liver." Archives of Biochemistry and Biophysics, vol. 400, no. 1, 2002, pp. 48-53.
Maia L, Mira L. Xanthine oxidase and aldehyde oxidase: a simple procedure for the simultaneous purification from rat liver. Arch Biochem Biophys. 2002;400(1):48-53.
Maia, L., & Mira, L. (2002). Xanthine oxidase and aldehyde oxidase: a simple procedure for the simultaneous purification from rat liver. Archives of Biochemistry and Biophysics, 400(1), 48-53.
Maia L, Mira L. Xanthine Oxidase and Aldehyde Oxidase: a Simple Procedure for the Simultaneous Purification From Rat Liver. Arch Biochem Biophys. 2002 Apr 1;400(1):48-53. PubMed PMID: 11913970.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Xanthine oxidase and aldehyde oxidase: a simple procedure for the simultaneous purification from rat liver. AU - Maia,Luisa, AU - Mira,Lurdes, PY - 2002/3/27/pubmed PY - 2002/5/7/medline PY - 2002/3/27/entrez SP - 48 EP - 53 JF - Archives of biochemistry and biophysics JO - Arch Biochem Biophys VL - 400 IS - 1 N2 - Aldehyde oxidase (AO) and xanthine oxidase (XO) are cytosolic enzymes that have been involved in some pathological conditions and play an important role in the biotransformation of drugs and xenobiotics. The increasing interest in these enzymes demands for a simple and rapid procedure for their purification. This paper describes for the first time a method that allows simultaneous purification of both enzymes from the same batch of rat livers. It involves few steps, is reproducible and offers high enzyme yields with high specific activities. The rat liver homogenate was fractionated by heat denaturation and by ammonium sulphate precipitation to give a crude extract containing both enzymes. This extract was chromatographed on an Hydroxyapatite column that completely separated AO from XO. Further purification of XO by anion exchange chromatography on a Q-Sepharose Fast Flow column resulted in a highly purified (1200-fold) preparation, with a specific activity of 3.64 U/mg and with a 20% yield. AO was purified about 1000-fold at a yield of 15%, with a specific activity of 3.48 U/mg, by affinity chromatography on Benzamidine-Sepharose 6B. The purified enzymes gave single bands of approximately 300 kDa on a polyacrylamide gel gradient electrophoresis and displayed the characteristic absorption spectra of highly purified enzymes. SN - 0003-9861 UR - https://www.unboundmedicine.com/medline/citation/11913970/Xanthine_oxidase_and_aldehyde_oxidase:_a_simple_procedure_for_the_simultaneous_purification_from_rat_liver_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0003986102927815 DB - PRIME DP - Unbound Medicine ER -