caa a22 4500 44537652X CHVBK 20180317143004.0 cr unu---uuuuu 170323e20110701xx s 000 0 eng 10.1007/s00702-011-0620-y doi (NATIONALLICENCE)springer-10.1007/s00702-011-0620-y Tyrosine 381 in E. coli copper amine oxidase influences substrate specificity [Elektronische Daten] [Christian Kurtis, Peter Knowles, Mark Parsons, Thembaninkosi Gaule, Simon Phillips, Michael McPherson] Copper amine oxidases are important for the metabolism of a range of biogenic amines. Here, we focus on substrate specificity in the E. coli copper amine oxidase (ECAO) and specifically the role of Tyr 381. This residue, and its equivalent, in other copper amine oxidases has been referred to as a "gating” residue able to move position depending upon the presence or absence of amine substrate. The position of this residue suggests a role in substrate selectivity. We have compared the properties of two variant forms of ECAO, Y381F and Y381A, with wild-type enzyme by steady-state kinetics of oxidation of a number of amine substrates, modes of inhibitor interactions and X-ray structure determination. Y381F displays a similar catalytic efficiency to wild type against the preferred substrate β-phenylethylamine. In both cases oxidation of the alternative aromatic amine substrate benzylamine is relatively poor, although Y381F represents an efficient benzylamine oxidase. By contrast, Y381A performed poorly against both aromatic substrates predominantly due to an increased K M which we propose is due to the lack of an aromatic residue to orient substrate towards the TPQ and active site base. These results are supported by different behaviour of Y381A to inhibition with 2-hydrazinopyridine. We also report on methylamine turnover by the three enzymes. We propose that Y381, together with another residue Y387, may be considered of critical importance for the substrate selectivity of ECAO, through stacking or hydrophobic interactions with substrate. Springer-Verlag, 2011 Escherichia coli nationallicence Copper amine oxidase nationallicence Trihydroxyphenyalanine quinine nationallicence β-Phenylethylamine nationallicence Benzylamine nationallicence Methylamine nationallicence Substrate specificity nationallicence 2HP : 2-Hydrazinopyridine nationallicence TCP : Tranylcypromine nationallicence ECAO : E. coli copper amine oxidase nationallicence TPQ : 2,4,5, Trihydroxyphenylalanine quinone nationallicence Kurtis Christian Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut Knowles Peter Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut Parsons Mark Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut Gaule Thembaninkosi Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut Phillips Simon Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut McPherson Michael Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut Journal of Neural Transmission Springer Vienna 118/7(2011-07-01), 1043-1053 0300-9564 118:7<1043 2011 118 702 https://doi.org/10.1007/s00702-011-0620-y text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s00702-011-0620-y text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Kurtis Christian Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut NATIONALLICENCE 700 1- Knowles Peter Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut NATIONALLICENCE 700 1- Parsons Mark Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut NATIONALLICENCE 700 1- Gaule Thembaninkosi Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut NATIONALLICENCE 700 1- Phillips Simon Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut NATIONALLICENCE 700 1- McPherson Michael Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, LS2 9JT, Leeds, UK aut NATIONALLICENCE 773 0- Journal of Neural Transmission Springer Vienna 118/7(2011-07-01), 1043-1053 0300-9564 118:7<1043 2011 118 702 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer