Cloning and Sequencing of Phenylethylamine Oxidase from Arthrobacter globiformis and Implication of Tyr-382 as the Precursor to Its Covalently Bound Quinone Cofactor

Tanizawa, Katsuyuki; Matsuzaki, Ryuichi; Shimizu, Eiichi; Yorifuji, Tohru; Fukui, Tsuguya

doi:10.1006/bbrc.1994.1343

Cited by 68 publications

(63 citation statements)

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“…Elucidation of the 1,4-diamin0-2-butynebinding site and its consensus sequence is now in progress. When the amino acid sequence near the active site, towards, the amino terminus, was compared for amine oxidases from various sources [23,[26][27][28][29][30][31][32], as shown in Fig. 9, some similarity around the position of the 1,4-diamino-2-butyne-binding site of A. niger AO-I was found.…”

Section: 4 1 E T I a D Y T G K P T T I P G A V A I F E R Y A G P E mentioning

confidence: 99%

Active‐Site Covalent Modifications of Quinoprotein Amine Oxidases from Aspergillus niger

Frébort¹,

Peč

Luhová

et al. 1994

European Journal of Biochemistry

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Interactions of two distinct quinoprotein amine oxidases from Aspergillus niger, AO-I and AO-11, with active-site covalent modifiers have been investigated. Both enzymes are inhibited similarly by phenylhydrazine or p-nitrophenylhydrazine, forming an orange Schiff base with a carbonyl group of topaquinone cofactor. Modification of histidyl and tyrosyl residues by diethylpyrocarbonate and sulfhydryl groups by 5,5'-dithio-bis-(2-nitrobenzoic acid) and 4-chloro-7-nitrobenzo-2-oxa-l,3-diazole have been described. A substrate analog, 1,4-diamino-2-butyne, was found to function as a mechanism-based inhibitor. It shows both substrate saturation kinetics and time-dependent irreversible inhibition caused by formation of pynole bound to the active site. The pyrrole formation was confirmed spectrophotometrically by reaction with Ehrlich's reagent at 525 nm. Inhibition by 1,4-diamino-2-butyne produces a new maximum in the absorption spectra of AO-I and AO-I1 at 310 nm and 306 nm, respectively. Inactivated AO-I was digested by proteases ; labeled peptides were purified by C18 HPLC and sequenced by Edman degradation. Data reveal the evidence that 1P-diamino-2-butyne reacts with the &-amino group of the Lys356 residue in the sequence Lys-Met-Pro-AsnAla of Aspergillus niger amine oxidase AO-I.Amine oxidase of Aspergillus niger is a copper-carbonyl enzyme, which was found in the mid 1960s, purified and crystallized [l, 21. Copper and an undetermined carbonyl cofactor were found to function in the catalytic cycle [2-41. Nowadays, the importance of the fungal amine oxidase has risen again because the enzyme can be applied to monitor histamine contents in fishery products [5]. A massive production of the enzyme on the industrial scale has increased current interest.Recently, we have found two amine oxidases (EC 1.4.3.6), named AO-I and AO-11, in Aspergillus niger mycelia grown on n-butylamine medium [6, 71. AO-I is the enzyme previously reported, consisting of two 75-kDa subunits, while AO-I1 is a novel enzyme of 80 kDa. These amine oxidases do not show big differences in their peptide maps, but differ in their N-terminals. They also do not show any remarkable difference in substrate specificity or sensitivity to inhibitors. The cofactors of both enzymes give a typical reaction with p-nitrophenylhydrazine [6, 71 like other topaquiCorrespondence to I. Frkbort, Laboratory of Applied Microbiology, Faculty of Agriculture, Yarnaguchi University, 1 677-1 Yoshida, Yamaguchi, Japan 753Abbreviations. AO-I and AO-11, Aspergillus niger amine oxidases ; Nbs,, 5,5'-dithio-bis-(2-nitrobenzoic acid) ; buffer A, 0.01 M potassium phosphate pH 7.0.Enzymes. Amine oxidase, arnine:O, oxidoreductase (dearninating) (copper containing) (EC 1.4.3.6) ; peroxidase from horseradish (EC 1.11.1.7); pronase P from Streptomyces griseus (EC 3.4.24.31); thermolysin from Bacillus themolyticus rokko (EC 3.4.24.27 It is very probable that AO-I was purified and crystallized in earlier studies but fractions containing AO-I1 were not purified further because of its lower...

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Section: 4 1 E T I a D Y T G K P T T I P G A V A I F E R Y A G P E mentioning

confidence: 99%

Active‐Site Covalent Modifications of Quinoprotein Amine Oxidases from Aspergillus niger

Frébort¹,

Peč

Luhová

et al. 1994

European Journal of Biochemistry

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“…Thus, Asn-Tyr-(Asp/Glu) is the consensus TPQ site in all copper amine oxidases, with an additional Tyr located toward the C terminus of Asp/Glu in most cases . It is evident that Tyr-382 in the conserved sequence of phenylethylamine oxidase is essential as the precursor to TPQ Matsuzaki et al, 1994). However, the significance of the conservation of the other residues remains unknown, except for the finding that Glu-406 in the conserved sequence of the yeast copper amine oxidase does not play a role in the TPQ biogenesis (Cai and Klinman, 1994a).…”

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confidence: 87%

“…Purification of Wild-type and Mutant Enzymes-E. coli JM109 cells carrying pUTH or one of the mutant plasmids were grown at 37°C in a copper-depleted medium (Matsuzaki et al, 1994;Choi et al, 1995) supplemented with 50 g/ml sodium ampicillin. Cells were cultivated until the cell density reached A 600 ϭ 0.8 (about 3 h); then 50 M isopropyl-1-thio-␤-D-galactopyranoside was added and the bacteria were further cultivated at 30°C for 6 h. The wild-type and mutant enzymes were purified to homogeneity in the copper-free form using the procedure reported previously (Choi et al, 1995).…”

Section: Sequencementioning

confidence: 99%

“…This shows that the inactive enzyme is a precursor protein that can form TPQ through a self-processing mechanism, as first demonstrated with phenylethylamine oxidase from the same bacterial species (Matsuzaki et al, 1994). To elucidate the role(s) of the consensus sequence in the TPQ biogenesis, we have prepared a series of histamine oxidase proteins site specifically mutated at residues 401-404 in the conserved Asn-Tyr-Asp-Tyr sequence.…”

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confidence: 99%

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Role of Conserved Asn-Tyr-Asp-Tyr Sequence in Bacterial Copper/2,4,5-Trihydroxyphenylalanyl Quinone-containing Histamine Oxidase

Choi

Matsuzaki

Suzuki

et al. 1996

Journal of Biological Chemistry

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Copper amine oxidase contains a covalently bound quinonoid cofactor, 2,4,5-trihydroxyphenylalanyl quinone (TPQ), which is synthesized by post-translational modification of a specific tyrosyl residue occurring in the highly conserved sequence, Asn-Tyr-(Asp/Glu)-Tyr. To elucidate the role(s) of the conserved sequence in the biogenesis of TPQ, each of the corresponding residues at positions 401-404 in the recombinant histamine oxidase from Arthrobacter globiformis has been replaced with other amino acids by site-directed mutagenesis. When Asn-401 was changed to Asp or Gln, the rate of TPQ formation by copper-dependent self-processing was 10 3 -to 10 4 -fold slower than in the wild-type enzyme. When Tyr-402 was replaced by Phe, TPQ was not formed at all, showing that Tyr-402 is essential as the precursor to TPQ. In contrast, Asp-403 could be replaced by Glu without changes in the rate of TPQ formation, whereas its replacement by Asn led to a marked decrease. Furthermore, when Tyr-404 was changed to Phe, TPQ was formed swiftly on incubation with copper ions, but the TPQ enzyme exhibited very low activity with altered substrate specificity. These results collectively indicate that a very rigorous structural motif is required for efficient formation of TPQ and for the catalytic activity in the active site of copper amine oxidases.

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“…Recent progress in the study of TOPA-containing amine oxidases has provided evidence for cofactor formation via enzyme-bound copper in a post-translational self-processing event [1][2][3]. When amino acid sequences from four eukaryotic amine oxidases were aligned, a C-terminal motif consisting of the conserved positioning of various histidines in relation to the cofactor site became apparent [4].…”

Section: Introductionmentioning

confidence: 99%

Limited proteolysis of Hansenula polymorpha yeast amine oxidase: isolation of a C‐terminal fragment containing both a copper and quino‐cofactor

Plastino

Klinman

1995

FEBS Letters

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Limited proteolysis of recombinant Hansenula polymorpha yeast amino oxidase produces a 48 kDa fragment which corresponds to the C-terminal two-thirds of the protein. The fragment contains both TOPA (2,4,S-trihydroxyphenylalanine) and copper, as well as the histidine ligands implicated in copper binding. The fragment is proposed to be the domain responsible for cofactor production in yeast amine oxidase.

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Cloning and Sequencing of Phenylethylamine Oxidase from Arthrobacter globiformis and Implication of Tyr-382 as the Precursor to Its Covalently Bound Quinone Cofactor

Cited by 68 publications

References 0 publications

Active‐Site Covalent Modifications of Quinoprotein Amine Oxidases from Aspergillus niger

Active‐Site Covalent Modifications of Quinoprotein Amine Oxidases from Aspergillus niger

Role of Conserved Asn-Tyr-Asp-Tyr Sequence in Bacterial Copper/2,4,5-Trihydroxyphenylalanyl Quinone-containing Histamine Oxidase

Limited proteolysis of Hansenula polymorpha yeast amine oxidase: isolation of a C‐terminal fragment containing both a copper and quino‐cofactor

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