[2022 AWARD WINNING PAPER] Red-spotted grouper Epinephelus akaara blood L-amino acid oxidase utilizes the substrates in plasma

Fish Sci (2022) 88:635–643
DOI: https://doi.org/10.1007/s12562-022-01617-x

Authors

Yoichiro Kitani

Abstract

L-amino acid oxidases (LAOs) oxidatively deaminate L-amino acids and generate H₂O₂. Recently, a novel LAO was isolated from the red-spotted grouper Epinephelus akaara serum (EaLAO) and it showed antibacterial activity via H₂O₂. The enzymatic activity of EaLAO was normally suppressed in blood and was activated by mixing it with seawater. This phenomenon suggested that EaLAO may stand by as an inactive form in the body, and it functions as an antibacterial protein at the bleeding trauma sites. For pathogen protection by EaLAO at the wound site, instant H₂O₂ production is essential. In this work, the substrate source of EaLAO and the H₂O₂-generation potential of the grouper plasma were studied. Mixing plasma with seawater or cations generated 0.26–0.48 mM of H₂O₂ except for the addition of L-amino acid substrates. The amino acid quantification of plasma via fluorescent HPLC showed that the alanine was dominantly consumed after being mixed with seawater. The V_max, K_m, and turnover number of EaLAO with L-alanine at 25 °C in seawater were 1618 μU/reaction, 86.6 mM, and 10.6 s⁻¹, respectively. Furthermore, Vibrio harveyi was 84% killed by the plasma–seawater mixture. These results suggested that L-alanine may be the primary substrate of EaLAO in plasma.

Keywords

L-amino acid oxidase (LAO) / Epinephelus akaara / Substrate amino acids / Plasma

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