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Purification and characterisation of monoamine oxidase from Avena sativa

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Abstract

FAD-containing monoamine oxidase (MAO; EC 1.4.3.4) oxidises monoamines to their corresponding aldehydes, H2O2, and NH3. It has been purified to homogeneity in mammals, but to our knowledge, there have been no reports of the enzyme in plants. MAO activity was detected in Avena sativa seedlings during germination using benzylamine as substrate. The enzyme was purified to homogeneity (as assessed by native PAGE) by Sephadex G-25, DEAE Sephacel, hydroxyapatite, Mono Q, and TSK-GEL column chromatographies. The molecular mass estimated by gel filtration using the TSK-GEL column was 220 kDa. SDS-PAGE yielded four distinct protein bands of 78, 58, 55, and 32 kDa molecular masses. The pI value of the enzyme was 6.3. The enzyme showed high substrate specificity for an endogenous amine, phenethylamine, which was oxidised to phenylacetaldehde, but not for ethylamine, propylamine, butylamine, pentylamine, dopamine, serotonin, tryptamine, or tyramine. The K m values for benzylamine and phenethylamine were 2.7 × 10−4 and 7.1 × 10−4 M, respectively. Enzyme activity was not inhibited by pargyline, clorgyline, semicarbazide, or Na-diethyldithiocarbamate. Benzaldehyde, the product of benzylamine oxidation, exhibited strong competitive inhibition of enzyme activity with a Ki of 3 μM. FAD was identified by ODS-column chromatography as an enzyme cofactor. The enzyme contained 2 mol of FAD per 220,000 g of enzyme.

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Authors and Affiliations

  1. Department of Bio-GeoSciences, Osaka City University, Osaka, 558-8585, Japan

    Yong-Ming Zhang, Jeyanthi Rebecca Livingstone & Eiji Hirasawa

  2. College of Life Science and Technology, Inner Mongolia Normal University, Hohhot, 010022, Inner Mongolia, China

    Yong-Ming Zhang

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  1. Yong-Ming Zhang
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  2. Jeyanthi Rebecca Livingstone
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  3. Eiji Hirasawa
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Correspondence to Eiji Hirasawa.

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Communicated by Z.-L. Zhang.

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Zhang, YM., Livingstone, J.R. & Hirasawa, E. Purification and characterisation of monoamine oxidase from Avena sativa . Acta Physiol Plant 34, 1411–1419 (2012). https://doi.org/10.1007/s11738-012-0939-5

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  • DOI: https://doi.org/10.1007/s11738-012-0939-5

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