Tedeschi Gabriella, Pollegioni Loredano, Negri Armando
Dipartimento di Patologia Animale, Igiene e Sanità Pubblica Veterinaria - sez. Biochimica, Università degli Studi di Milano, Milano, Italy.
Methods Mol Biol. 2012;794:381-95. doi: 10.1007/978-1-61779-331-8_26.
D-Amino acid oxidase and D-aspartate oxidase are two well-known FAD-containing flavooxidases that catalyze the same reaction (the oxidative deamination) on different D-amino acids. D-aspartate oxidase is specific for acidic D-amino acids (i.e., D-aspartate and D-glutamate) and D-amino acid oxidase is active on neutral and polar D-amino acids (a low activity is also detected on basic D-amino acids). The assay of these flavoenzymes is of utmost importance in different fields because D-amino acids are common constituents of bacterial cell walls, are present in foods and because free D-serine and D-aspartic acid were identified in brain and peripheral tissues of mammals. In this chapter, we report on the most used methods employed to assay the activity of D-amino acid oxidase and D-aspartate oxidase. Interestingly, their activity can be followed using different assays, namely D-amino acid or oxygen consumption, α-keto acid or ammonia production, or using artificial dyes as final indicator of the flavin redox reaction.
D-氨基酸氧化酶和D-天冬氨酸氧化酶是两种著名的含黄素腺嘌呤二核苷酸（FAD）的黄素氧化酶，它们对不同的D-氨基酸催化相同的反应（氧化脱氨反应）。D-天冬氨酸氧化酶对酸性D-氨基酸（即D-天冬氨酸和D-谷氨酸）具有特异性，而D-氨基酸氧化酶对中性和极性D-氨基酸具有活性（在碱性D-氨基酸上也检测到低活性）。这些黄素酶的测定在不同领域至关重要，因为D-氨基酸是细菌细胞壁的常见成分，存在于食物中，还因为在哺乳动物的脑和外周组织中发现了游离的D-丝氨酸和D-天冬氨酸。在本章中，我们报告了用于测定D-氨基酸氧化酶和D-天冬氨酸氧化酶活性的最常用方法。有趣的是，可以使用不同的测定方法来跟踪它们的活性，即D-氨基酸或氧气消耗、α-酮酸或氨的产生，或者使用人工染料作为黄素氧化还原反应的最终指示剂。
