Biomaterial Database

A new purification procedure for fumarase based of affinity chromatography. Isolation and characterization of pig-liver fumarase. 1977

S Beeckmans, and L Kanarek

A new procedure has been developed for the isolation of fumarase (EC 4.2.1.2). It is described for the purification of pigheart and liver enzyme. Pyromellitic acid has been covalently coupled to Sepharose-4B with diaminopropanol as spacer arr. When a dialysed 0.55 saturated ammonium sulphate precipitate is applied to the column, in Tris-acetate buffer, pH 7.3, fumarase remains quantitatively bound. It is eluted by competition, together with a few other proteins, by the natural product L-malate. Malate is removed from the eluate by dialysis. After this highly efficient purification step the enzyme is very easily crystallized. The final yield by 67% for both pig heart and liver preparations. The specific activity of fumarase purified from both tissues is found to be the same. Polyacrylamide gel electrophoresis in dodecylsulphate shows one single band corresponding with a subunit molecular weightof 48500. A single band is also obtained by electrophoresis in acid urea. This new procedure based on biospecific affinity chromatography allows a fast and easy preparation of gram quantities of fumarase.

UI	MeSH Term	Description	Entries
D007461	Iodoacetates	Iodinated derivatives of acetic acid. Iodoacetates are commonly used as alkylating sulfhydryl reagents and enzyme inhibitors in biochemical research.	Iodoacetic Acids,Acids, Iodoacetic
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D008024	Ligands	A molecule that binds to another molecule, used especially to refer to a small molecule that binds specifically to a larger molecule, e.g., an antigen binding to an antibody, a hormone or neurotransmitter binding to a receptor, or a substrate or allosteric effector binding to an enzyme. Ligands are also molecules that donate or accept a pair of electrons to form a coordinate covalent bond with the central metal atom of a coordination complex. (From Dorland, 27th ed)	Ligand
D008099	Liver	A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances.	Livers
D009206	Myocardium	The muscle tissue of the HEART. It is composed of striated, involuntary muscle cells (MYOCYTES, CARDIAC) connected to form the contractile pump to generate blood flow.	Muscle, Cardiac,Muscle, Heart,Cardiac Muscle,Myocardia,Cardiac Muscles,Heart Muscle,Heart Muscles,Muscles, Cardiac,Muscles, Heart
D009928	Organ Specificity	Characteristic restricted to a particular organ of the body, such as a cell type, metabolic response or expression of a particular protein or antigen.	Tissue Specificity,Organ Specificities,Specificities, Organ,Specificities, Tissue,Specificity, Organ,Specificity, Tissue,Tissue Specificities
D010071	Oxaloacetates	Derivatives of OXALOACETIC ACID. Included under this heading are a broad variety of acid forms, salts, esters, and amides that include a 2-keto-1,4-carboxy aliphatic structure.	Ketosuccinates,Oxosuccinates,Oxaloacetic Acids
D002846	Chromatography, Affinity	A chromatographic technique that utilizes the ability of biological molecules, often ANTIBODIES, to bind to certain ligands specifically and reversibly. It is used in protein biochemistry. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)	Chromatography, Bioaffinity,Immunochromatography,Affinity Chromatography,Bioaffinity Chromatography
D005649	Fumarate Hydratase	An enzyme that catalyzes the reversible hydration of fumaric acid to yield L-malic acid. It is one of the citric acid cycle enzymes. EC 4.2.1.2.	Fumarase,Hydratase, Fumarate
D000596	Amino Acids	Organic compounds that generally contain an amino (-NH2) and a carboxyl (-COOH) group. Twenty alpha-amino acids are the subunits which are polymerized to form proteins.	Amino Acid,Acid, Amino,Acids, Amino