Binding of [14C] allylamine to isolated mitochondria from rat heart and aorta.
This study demonstrates specific and saturable binding of [14C] allylamine to mitochondria derived from rat aorta and heart. Specific binding is linear with respect to mitochondrial concentration and has a pH optimum of 7.0. Saturation isotherms reveal anomalous kinetics of specific binding on heart mitochondria with a high affinity site (KD 16 nM) and a lower affinity site (KD 80 nM); Scatchard plots have a common intercept. Exhaustive flow dialysis in the presence of SDS demonstrates that as much as 23.5% of bound radioactive moieties in aorta mitochondria are covalently bound, and as much as 42.6% are covalently bound in heart mitochondria. Hydrolysis of heart mitochondria with phospholipase C markedly enhances saturation of [14C] allylamine, and greatly increases the quantity of covalently bound radioactive ligand. Phospholipase C hydrolysis of heart mitochondria increased monoamine oxidase B activities and unmasked a small amount of benzylamine oxidase activity, whereas hydrolysis of mitochondria with phospholipases A2 and D diminish MAO-B activity. The monoamine oxidase B inhibitor, deprenyl, significantly reduced both specific and covalent binding of the 14C-activity from [14C] allylamine to phospholipase hydrolyzed mitochondria. The benzylamine oxidase inhibitor, phenelzine, significantly decreased specific binding but had no effect on the degree of covalent binding of [14C] allylamine to phospholipase C hydrolyzed mitochondria. The benzylamine oxidase inhibitor, semicarbazide, had no effect in inhibiting [14C] allylamine binding. Covalent binding of 14C-moiety from [14C] allylamine to mitochondria--which express specific binding sites for the [14C] allylamine--and inhibition of binding by monoamine oxidase inhibitors, suggest the formation of highly reactive intermediates.
D000499:Allylamine / Q000378:metabolism*; D000588:Amines / Q000378:metabolism*; D000818:Animals; D001011:Aorta / Q000378:metabolism*; D001665:Binding Sites; D002250:Carbon Radioisotopes; D066298:In Vitro Techniques; D008297:Male; D008929:Mitochondria, Heart / Q000378:metabolism*; D008931:Mitochondria, Muscle / Q000378:metabolism*; D008995:Monoamine Oxidase / Q000032:analysis; D010624:Phenelzine / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D012642:Selegiline / Q000494:pharmacology; D010738:Type C Phospholipases / Q000494:pharmacology
D000588:Amines; D002250:Carbon Radioisotopes; D012642:Selegiline; D000499:Allylamine; D008995:Monoamine Oxidase; D010738:Type C Phospholipases; D010624:Phenelzine