Effect of phosphate ion on the activity of bovine plasma amine oxidase.
The system bovine plasma amine oxidase-polyamine-phosphate ion was investigated by activity measurements and 31P NMR spectroscopy. Lineweaver-Burk plots showed that phosphate ion, under physiological conditions, is an apparent competitive inhibitor of bovine plasma amine oxidase. While NMR measurements of the T1 of 31P do not suggest the binding of phosphate to/or near the paramagnetic Cu(II) sites of bovine plasma amine oxidase, the chemical shift dependence of 31P on spermidine concentration indicates the formation of a spermidine-phosphate complex. The value of the dissociation constant of this complex was found 18.5 +/- 1.4 mM, at pH 7.2, by NMR, in good agreement with the value 17.0 +/- 0.8 mM calculated from activity measurements, assuming the enzyme activity is proportional to the free amine concentration, under second order conditions. Our data suggest that the decrease of the free spermidine, due to the binding of phosphate ion, is responsible of the observed inhibition of bovine plasma amine oxidase.
Corazza
A
A
Department of Biological Chemistry, University of Padova, Italy.
Stevanato
R
R
Di Paolo
M L
ML
Scarpa
M
M
Mondovì
B
B
Rigo
A
A
D006631:Amine Oxidase (Copper-Containing); D000818:Animals; D002417:Cattle; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008433:Mathematics; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000097:blood*; D010710:Phosphates / Q000494:pharmacology*; D010758:Phosphorus; D013095:Spermidine / Q000378:metabolism
D010710:Phosphates; D010758:Phosphorus; D006631:Amine Oxidase (Copper-Containing); D000587:Oxidoreductases Acting on CH-NH Group Donors; D013095:Spermidine