Copper amine oxidase from Hansenula polymorpha: the crystal structure determined at 2.4 å resolution reveals the active conformation

Li, Rongbao; Klinman, Judith P.; Mathews, F.S.

doi:10.1016/s0969-2126(98)00033-1

Cited by 183 publications

(260 citation statements)

References 43 publications

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“…HPAO Crystallization-Crystallization was performed as previously reported (13 mg/ml HPAO in 8.0 -9.5% polyethylene glycol 8000, 0.28 -0.30 M phosphate at pH 6.0 at 293 K) with the exception of the use of hanging drop crystallization rather than sitting drop (34). The ratio of protein to well solution in the hanging drops was 1:1, with volumes of 2.5 l, giving a total volume of 5.0 l. The best diffracting crystals were cube-shaped with dimensions of 40 ϫ 40 ϫ 30 m and grew within 3 days.…”

Section: Methodsmentioning

confidence: 99%

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Exploring Molecular Oxygen Pathways in Hansenula polymorpha Copper-containing Amine Oxidase

Johnson

Cohen

Welford

et al. 2007

Journal of Biological Chemistry

111

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The accessibility of large substrates to buried enzymatic active sites is dependent upon the utilization of proteinaceous channels. The necessity of these channels in the case of small substrates is questionable because diffusion through the protein matrix is often assumed. Copper amine oxidases contain a buried protein-derived quinone cofactor and a mononuclear copper center that catalyze the conversion of two substrates, primary amines and molecular oxygen, to aldehydes and hydrogen peroxide, respectively. The nature of molecular oxygen migration to the active site in the enzyme from Hansenula polymorpha is explored using a combination of kinetic, x-ray crystallographic, and computational approaches. A crystal structure of H. polymorpha amine oxidase in complex with xenon gas, which serves as an experimental probe for molecular oxygen binding sites, reveals buried regions of the enzyme suitable for transient molecular oxygen occupation. Calculated O 2 free energy maps using copper amine oxidase crystal structures in the absence of xenon correspond well with later experimentally observed xenon sites in these systems, and allow the visualization of O 2 migration routes of differing probabilities within the protein matrix. Site-directed mutagenesis designed to block individual routes has little effect on overall k cat /K m (O 2 ), supporting multiple dynamic pathways for molecular oxygen to reach the active site.Copper amine oxidases are ubiquitous copper containing enzymes that oxidize primary amines to aldehydes through the reduction of molecular oxygen to hydrogen peroxide. CAO 2 catalysis is dependent upon the protein-derived cofactor 2,4,5-trihydroxyphenylalaninequinone (TPQ). The TPQ is derived from an endogenous tyrosine through a self-catalytic process requiring only molecular oxygen and Cu(II) (see Fig. 1a) (1).Hansenula polymorpha 3 amine oxidase is the eukaryotic CAO that has been kinetically characterized in the most detail (2-7). HPAO follows a Bi Bi ping-pong reaction mechanism that can be expressed as two half-reactions, reductive and oxidative (see Fig. 1b). In the reductive half-reaction the enzyme oxidizes a primary amine to an aldehyde, generating the 2e Ϫ reduced aminoquinol form of the cofactor. In the subsequent oxidative half-reaction molecular oxygen is reduced to hydrogen peroxide via cofactor reoxidation to TPQ. Biochemical studies from several different laboratories have led to mechanistic proposals for the catalytic cycle of CAOs (8, 9). These studies have given significant insight into the mechanism for the reductive half-reaction (10). However, the details surrounding the activation of molecular oxygen both in terms of the biogenesis of the TPQ (11, 12) and of the catalytic oxidative half-reaction, remain the subject of intense study (13-16). The utilization of copper as a redox center has been the focus of recent controversy. Because CAOs contain a copper ion in their active site, chemical intuition suggests Cu(I) as the O 2 -activating species to give Cu(II)-superoxide (17). Upon a...

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Section: Methodsmentioning

confidence: 99%

“…2) (34,35). The inland lake is conserved at the dimer interface of CAOs and has been suggested as the entry point for O 2 (35).…”

mentioning

confidence: 99%

Exploring Molecular Oxygen Pathways in Hansenula polymorpha Copper-containing Amine Oxidase

Johnson

Cohen

Welford

et al. 2007

Journal of Biological Chemistry

111

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“…The three-dimensional structure of CuAOs from Escherichia coli (PDB code 1oac; Parsons et al, 1995), Pisum sativum (PDB code 1ksi; Kumar et al, 1996), Arthobacter globiformis (PDB code 1av4; Wilce et al, 1997) and Hanensula polymorpha (PDB code 1a2v; Li et al, 1998) and of the lysyl oxidase from Pichia pastoris (PDB code 1n9e; Duff et al, 2003) have been reported. They all share a similar fold and contain three domains called D2, D3 and D4.…”

Section: Introductionmentioning

confidence: 99%

Crystallization of a truncated soluble human semicarbazide-sensitive amine oxidase

et al. 2005

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Human semicarbazide-sensitive amine oxidase (SSAO) is a homodimeric copper-containing monoamine oxidase that occurs in both a membrane-bound and a soluble form. SSAO is also known as vascular adhesion protein-1 (VAP-1). A truncated soluble form of human SSAO (comprising residues 29±763) was expressed in human embryonic kidney 293 cells and puri®ed to homogeneity. Tetragonal crystals were obtained and a data set extending to 2.5 A Ê was collected. The crystals are merohedrally twinned and the estimation of the twinning fraction was complicated by pseudo-symmetry and the anisotropic character of the crystals. Using a recently developed method for twinning detection that is insensitive to phenomena such as anisotropy or pseudosymmetry [Padilla & Yeates (2003), Acta Cryst. D59, 1124±1130], the twinning fraction was estimated to be 0.3. The structure was eventually solved by molecular replacement in space group P4 3 .

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“…As summarized in a recent review (3), x-ray crystal structures of CAOs have been determined for a number of enzymes from various sources, including bacteria (Escherichia coli (16) and Arthrobacter globiformis (AGAO) (17)(18)(19)), yeasts (Hansenula polymorpha (recently reclassified as Pichia angusta) (HPAO-1 and HPAO-2) (20,21) and Pichia pastoris (lysyl oxidase) (22)), a fungus (Aspergillus nidulans (23)), a plant (Pisum savitum (24)), a mammal (Bos taurus (25)), and Homo sapiens (diamine oxidase and vascular adhesion protein 1, which is identical to semicarbazide-sensitive amine oxidase (26 -28)). The active site structures of these enzymes, including the positions of TPQ and Cu(II), are highly conserved, suggesting that they have a common mechanism for single-turnover TPQ biogenesis and catalytic amine oxidation.…”

mentioning

confidence: 99%

Probing the Catalytic Mechanism of Copper Amine Oxidase from Arthrobacter globiformis with Halide Ions

Murakawa

Hamaguchi

Nakanishi

et al. 2015

Journal of Biological Chemistry

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Background:Copper amine oxidases catalyze amine oxidation using copper and a quinone cofactor. Results: Halides bind axially to the copper center, preventing the reduced cofactor from adopting an on-copper conformation. Conclusion:The cofactor undergoes large conformational changes during the catalytic reaction that enable transitions between different types of chemistry. Significance: Molecular details of cofactor movement have been unveiled based on structural and kinetic evidence.

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Copper amine oxidase from Hansenula polymorpha: the crystal structure determined at 2.4 å resolution reveals the active conformation

Cited by 183 publications

References 43 publications

Exploring Molecular Oxygen Pathways in Hansenula polymorpha Copper-containing Amine Oxidase

Exploring Molecular Oxygen Pathways in Hansenula polymorpha Copper-containing Amine Oxidase

Crystallization of a truncated soluble human semicarbazide-sensitive amine oxidase

Probing the Catalytic Mechanism of Copper Amine Oxidase from Arthrobacter globiformis with Halide Ions

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