1iqy: Difference between revisions

==Overview==

The role of the active site Cu(2+) of phenylethylamine oxidase from, Arthrobacter globiformis (AGAO) has been studied by substitution with, other divalent cations, where we were able to remove >99.5% of Cu(2+) from, the active site. The enzymes reconstituted with Co(2+) and Ni(2+) (Co- and, Ni-AGAO) exhibited 2.2 and 0.9% activities, respectively, of the original, Cu(2+)-enzyme (Cu-AGAO), but their K(m) values for amine substrate and, dioxygen were comparable. X-ray crystal structures of the Co- and Ni-AGAO, were solved at 2.0-1.8 A resolution. These structures revealed changes in, the metal coordination environment when compared to that of Cu-AGAO., However, the hydrogen-bonding network around the active site involving, metal-coordinating and noncoordinating water molecules was preserved. Upon, anaerobic mixing of the Cu-, Co-, and Ni-AGAO with amine substrate, the, 480 nm absorption band characteristic of the oxidized form of the, topaquinone cofactor (TPQ(ox)) disappeared rapidly (< 6 ms), yielding the, aminoresorcinol form of the reduced cofactor (TPQ(amr)). In contrast to, the substrate-reduced Cu-AGAO, the semiquinone radical (TPQ(sq)) was not, detected in Co- and Ni-AGAO. Further, in the latter, TPQ(amr) reacted, reversibly with the product aldehyde to form a species with a lambda(max), at around 350 nm that was assigned as the neutral form of the product, Schiff base (TPQ(pim)). Introduction of dioxygen to the substrate-reduced, Co- and Ni-AGAO resulted in the formation of a TPQ-related intermediate, absorbing at around 360 nm, which was assigned to the neutral iminoquinone, form of the 2e(-)-oxidized cofactor (TPQ(imq)) and which decayed, concomitantly with the generation of TPQ(ox). The rate of TPQ(imq), formation and its subsequent decay in Co- and Ni-AGAO was slow when, compared to those of the corresponding reactions in Cu-AGAO. The low, catalytic activities of the metal-substituted enzymes are due to the, impaired efficiencies of the oxidative half-reaction in the catalytic, cycle of amine oxidation. On the basis of these results, we propose that, the native Cu(2+) ion has essential roles such as catalyzing the electron, transfer between TPQ(amr) and dioxygen, in part by providing a binding, site for 1e(-)- and 2e(-)-reduced dioxygen species to be efficiently, protonated and released and also preventing the back reaction between the, product aldehyde and TPQ(amr).

==About this Structure==

1IQY is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Arthrobacter_globiformis Arthrobacter globiformis] with NI as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Amine_oxidase_(copper-containing) Amine oxidase (copper-containing)], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.4.3.6 1.4.3.6] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1IQY OCA].

 

Role of copper ion in bacterial copper amine oxidase: spectroscopic and crystallographic studies of metal-substituted enzymes., Kishishita S, Okajima T, Kim M, Yamaguchi H, Hirota S, Suzuki S, Kuroda S, Tanizawa K, Mure M, J Am Chem Soc. 2003 Jan 29;125(4):1041-55. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=12537504 12537504]

[[Category: Single protein]]

[[Category: Hirota, S.]]

[[Category: Kim, M.]]

[[Category: Kishishita, S.]]

[[Category: Kuroda, S.]]

[[Category: Mure, M.]]

[[Category: Okajima, T.]]

[[Category: Tanizawa, K.]]

[[Category: Yamaguchi, H.]]