1-amino Acid Oxidase in Leukocytes: a Possible

-D-Amino acid oxidase has been identified within the granule fraction of human neutrophilic leukocytes. Leukocyte homogenates and purified kidney 1-amino acid oxidase can utilize either isolated D-amino acids or some species of bacteria as substrates for the generation of hydrogen peroxide. When linked to leukocyte myeloperoxidase in vitro, purified D-amino acid oxidase constitutes a system lethal for certain bacteria. It is proposed that leukocyte D-amino acid oxidase and myeloperoxidase constitute a biochemically specific system for the recognition and killing of certain microorganisms. Phagocytic leukocytes constitute a phylogenetically primitive system of resistance to microbial infection. Reduced to its simplest elements, this system must perform at least two integrated functions: the ingestion and the killing of microorganisms. The requirements for particle uptake by leukocytes are known in considerable detail." 2 The mechanisms of microbial killing are less well defined, although some weapons in the arsenal of the phagocytic leukocyte have been extensively studied.3' 4 A leukocyte microbicidal system consisting of the leukocyte enzyme myeloperoxidase, a halide, and hydrogen peroxide or a hydrogen peroxide-generating system has recently been described.5 The hypothesis that this system plays a role in the normal defense function of the human neutrophilic leukocyte is supported by the observation of impaired microbicidal activity in leukocytes genetically deficient in myeloperoxidase6 or in postphagocytic hydrogen peroxide generation.7' 8 Several considerations led us to predict the existence in leukocytes of a hydrogen peroxide-generating system linked to D-amino acid oxidase. D-Amino acids are an integral part of the cell walls of many bacterial species9 and have been found in the products of certain fungi, but have not been conclusively identified as a natural component of any mammalian tissues. Nevertheless, the activity of 1-amino acid oxidase is known to occur in certain mammalian tissues and to be abundant in the liver and kidney. This enzyme, in the presence of molecular oxygen and the appropriate D-amino acid, catalyzes the generation of a corresponding keto-acid and of hydrogen peroxide. This communication reports the finding of D-amino acid oxidase in the granule fraction of mammalian neutrophils. The enzyme utilized intact microorganisms as a substrate for hydrogen peroxide generation and could be linked with human myeloperoxidase in vitro to constitute a potent microbicidal system. Materials and Methods.-Preparation of enzymes: (a) D-Amino acid oxidase: Populations of either mixed leukocytes or lymphocytes were prepared from the heparinized venous blood of normal subjects and patients with hematologic diseases.'0 Neutrophil-rich