โ Scribed by Donald T. Sawyer; Tohru Tsuchiya; G.Susan Srivatsa
No coin nor oath required. For personal study only.
1st Intern. Conf. on Bioinorganic Chemistry -Session E inhibitors of bovine plasma and pig kidney amine oxidase [ 3,4] .
Cu(I1) complexes of 1 ,lO-phenanthroline (phen) and 2,2'-bipyridine (bipy) strongly inhibited both amine oxidases. Lysine, tyrosine, and salicylate complexes, as well as Cu*+(aq.), inhibited the plasma but not the kidney amine oxidase. No inhibition by the free ligands, by Cu(EDTA)*-, or bovine erythrocyte SOD was observed. Cu(phen)2(N03)2, Cu*+(aq.), and Cu(salicylate), displayed linear noncompetitive inhibition against amine substrates at saturating O2 concentrations with either pig kidney or beef plasma amine oxidase. Since neither the free ligands nor Cu*+(aq.) inhibit the kidney enzyme at concentrations where Cu(phen)2(N0s)2
and Cu(bipy),Clz are effective, the observed inhibitory action must be attributed to the complexes. Two plausible inhibition mechanisms are (1) dismutation of O,, which is an inter-. mediate in the reoxidation reaction; or (2) direct oxidation of Ered by the Cu(I1) complexes. Steric requirements for the second mechanism should be less restrictive than for mechanism (1) as outer-sphere electron transfer is possible over substantial distances, but 0, dismutation is an inner-sphere process that requires the Cu(I1) complex to penetrate to the O2 reduction site. Thus if 0, is tightly bound within the dioxygen-reduction site, its accessibility to Cu(I1) complexes of varying size and charge would be mechanistically important. On this basis the reactivity pattern we have observed can be rationalized: only those complexes with hydrophobic ligands and a net positive charge inhibited pig kidney amine oxidase, but the active site in the plasma enzyme may be much more accessible to external reagents. Nj-and SCN-are Cu(II)-specific inhibitors of several amine oxidases [ 1, S-71 . Ligand-to-metal charge-transfer bands are evident at -365 nm (E -3,000 M-' cm-') and -400 nm (E -6,000 M-' cm-') when SCN-and Na-, respectively, are added to beef plasma or pig kidney amine oxidase. The intensities and energies establish that the anions are equitorially coordinated.
Changes in the Cu(II) ligand field, as judged by absorption, circular dichroism and EPR spectroscopy, are consistent with a simple ligand substitution reaction. Preliminary NMR evidence indicates that Hz0 is the leaving group, as previously demonstrated for the pig plasma enzyme [6]. Depending on the amine concentration both anions are linear noncompetitive or linear uncompetitive inhibitors of beef plasma amine oxidase with O2 saturating; under the same conditions these anions were strictly linear uncompetitive inhibitors of the pig kidney enzyme. By determining in detail the inhibition mechanism of anions and the Cu(II) complexes, we should be able to learn much about the role(s) of copper in the amine oxidases.
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