title: Mode of inactivation of putrescine oxidase by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide or metal ions.
abstract: 1. Putrescine oxidase [EC 1.4.3.10] was partially inactivated by reaction with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), rapidly. The activity of the modified enzyme toward putrescine was 5.6% of that of the native enzyme. Moreover, the optimal pH and Km value of the modified enzyme for putrescine changed from 9.0 to 10.5 and from 0.23 to 3.4 mM, respectively. This modified enzyme showed activity toward monoamines such as n-butylamine, n-hexylamine, and n-octylamine, which are not substrates of the native enzyme. 2. Heavy metal ions inactivated putrescine oxidase completely. Hg2+ had the stronger ability to inactivate it. This inactivation was due to the dissociation of FAD from the enzyme molecule. The Fad-free enzyme had no ability to reassociate FAD, in contrast to the apoenzyme. The enzyme treated with Hg2+ showed characteristic behavior on SDS-polyacrylamide gel electrophoresis. 3. From the results of chemical modification it was deduced that a carboxyl group plays an important role in binding the substrate.
mesh_terms: D001051:Apoenzymes / Q000037:antagonists & inhibitors; D001665:Binding Sites; D002413:Cations, Divalent; D002942:Circular Dichroism; D005022:Ethyldimethylaminopropyl Carbodiimide / Q000494:pharmacology*; D005182:Flavin-Adenine Dinucleotide / Q000494:pharmacology; D007700:Kinetics; D008628:Mercury / Q000494:pharmacology; D008837:Micrococcus / Q000201:enzymology*; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000037:antagonists & inhibitors*; D011485:Protein Binding; D011487:Protein Conformation; D013379:Substrate Specificity
chemical_list: D001051:Apoenzymes; D002413:Cations, Divalent; D005182:Flavin-Adenine Dinucleotide; C022488:putrescine oxidase; D000587:Oxidoreductases Acting on CH-NH Group Donors; D008628:Mercury; D005022:Ethyldimethylaminopropyl Carbodiimide