Summary
Two distinct copper-quinoprotein amine oxidases (EC 1.4.3.6), AO-I and AO-II, were isolated from the soluble fraction of homogenized Aspergillus niger mycelia grown on butylamine medium. AO-I is a dimmer consisting of two 75 kDa subunits, while AO-II is a monomer of 80 kDa. Purified AO-I and AO-II show pink (490 nm) and yellow (420 nm) colors, respectively. The enzymes show comparable substrate specificity and sensitivity to inhibitors. Hexylamine, butylamine, benzylamine, tyramine and histamine are preferred substrates. Both enzymes are inhibited by substrate analogs, copper chelating agents, some alkaloids and carbonyl reagents. Enzymes labeled with p-nitrophenylhydrazine showed different absorption maxima at 465 and 440 nm for AO-I and AO-II, respectively, in neutral pH. These maxima were shifted to 585 nm under alkali conditions. Absorption and fluorescence spectra of synthesized model compound — topaquinone hydantoin p-nitrophenylhydrazone and labeled enzymes were in very good agreement, while PQQ p-nitrophenylhydrazone showed significant difference. After digestion by thermolysin, labeled cofactor containing peptides were purified using reverse phase HPLC and sequenced by Edman degradation, resulting typical topaquinone containing sequence, Asn-topa-Glu-Tyr, for AO-II and a noncofactor peptide, Val-Val-Ile-Glu-Pro, which contained topaquinone linked to γ-carboxyl of Glu, for AO-I. Cofactor structures were confirmed by mass spectrometry. N-Terminal amino acid sequences of AO-I and II were found to be different and peptide mapping showed some distinct patterns for both enzymes. A gene encoding AO-I has been cloned already and the second gene for AO-II is now under processing.
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© 1994 Birkhäuser Verlag Basel/Switzerland
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Frebort, I. et al. (1994). Two distinct quinoprotein amine oxidases are functioning in Aspergillus niger . In: Marino, G., Sannia, G., Bossa, F. (eds) Biochemistry of Vitamin B6 and PQQ. Advances in Life Sciences. Birkhäuser Basel. https://doi.org/10.1007/978-3-0348-7393-2_39
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DOI: https://doi.org/10.1007/978-3-0348-7393-2_39
Publisher Name: Birkhäuser Basel
Print ISBN: 978-3-0348-7395-6
Online ISBN: 978-3-0348-7393-2
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