Properties of cobalt-substituted bovine serum amine oxidase.
Half-copper-depleted and fully copper-depleted amine oxidase from bovine serum were reconstituted with either copper or cobalt. All samples were studied by high-sensitivity scanning calorimetry, by enzyme activity analysis, and by reactivity with phenylhydrazine. The calorimetric profile of the protein was strongly modified by the removal of a single Cu ion approximately to the same extent as by complete copper removal, in agreement with the loss of over 80% enzymic activity. The thermograms of metal-reconstituted species showed a marked similarity with that of the native enzyme, irrespective of whether copper or cobalt was present. Reactivity with phenylhydrazine and enzymic activity measurements showed that in cobalt-substituted amine oxidase the organic cofactor was reactive and the enzyme was catalytically competent, although kinetically less efficient. These observations agree both with previous findings on the protein half-site reactivity and with previous suggestions for a copper conformational role in bovine serum amine oxidase, namely of maintaining a functional conformation at the active site.
Department of Biochemical Sciences A. Rossi Fanelli, University of Rome La Sapienza, Italy.
E
E
Agostinelli
L
L
Morpurgo
C
C
Wang
A
A
Giartosio
B
B
Mondovì
D006631:Amine Oxidase (Copper-Containing); D000818:Animals; D002151:Calorimetry; D002417:Cattle; D003035:Cobalt / Q000737:chemistry* / Q000378:metabolism; D003300:Copper / Q000737:chemistry* / Q000378:metabolism; D004795:Enzyme Stability; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000097:blood / Q000737:chemistry / Q000378:metabolism*; D010659:Phenylhydrazines / Q000378:metabolism; D011487:Protein Conformation; D011489:Protein Denaturation; D013696:Temperature; D013816:Thermodynamics
D010659:Phenylhydrazines; C030299:phenylhydrazine; D003035:Cobalt; D003300:Copper; D006631:Amine Oxidase (Copper-Containing); D000587:Oxidoreductases Acting on CH-NH Group Donors