Purification and general properties of argininosuccinate lyase from jack bean, Canavalia ensiformis (L.)DC.
1. Argininosuccinate lyase (EC 4.3.2.1) from jack bean [Canavalia ensiformis (L.) DC] seeds was purified 532-fold from an acetone-butanol-dried powder. 2. The enzyme functions reversibly and exhibits maximum stability at 16 degrees . 3. At 16 degrees it has a half-life (t((1/2))) of 263min. 4. The enzyme is both cold-labile (t((1/2)) 131min. at 0 degrees ) and heat-inactivated (t((1/2)) 74min. at 38 degrees ); inactivation appears to be irreversible. 5. Treatment of the acetone-butanol-extracted powder with sodium dodecyl sulphate increased the sensitivity of the enzyme to temperature (t((1/2)) 70min. at 0 degrees ; t((1/2)) 23min. at 38 degrees ). 6. Addition, to the purified enzyme, of a fraction containing lipid from the seed increased the half-life to about 510min. at either 0 degrees or 38 degrees . 7. Arginine or homoarginine, and to a smaller extent some other amino acids or fumarate, protected the enzyme from cold-inactivation. 8. Reactivation attempts with both the cold- and heat-inactivated enzyme failed. 9. The K(m) value for argininosuccinate at pH7.5 is 1.3x10(-4). 10. The enzyme was inactivated completely within 15min. at 16 degrees by 0.5mm-p-hydroxymercuribenzoate, and subsequent exposure to 5mm-cysteine had no restorative effect.
D001120:Arginine; D001565:Benzoates; D055598:Chemical Phenomena; D002621:Chemistry; D002850:Chromatography, Gel; D003545:Cysteine; D003902:Detergents; D004355:Drug Stability; D005033:Ethylmaleimide; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008055:Lipids; D008190:Lyases / Q000302:isolation & purification; D008628:Mercury; D010944:Plants / Q000201:enzymology*; D013386:Succinates; D013431:Sulfates; D013696:Temperature; D013997:Time Factors
D001565:Benzoates; D003902:Detergents; D008055:Lipids; D013386:Succinates; D013431:Sulfates; D001120:Arginine; D008190:Lyases; D008628:Mercury; D003545:Cysteine; D005033:Ethylmaleimide