Roh J H, Suzuki H, Azakami H, Yamashita M, Murooka Y, Kumagai H
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
Biosci Biotechnol Biochem. 1994 Sep;58(9):1652-6. doi: 10.1271/bbb.58.1652.
The gene for monoamine oxidase (MAO) was cloned from an Escherichia coli genomic library and MAO was overproduced in the periplasmic space. The enzyme was purified to homogeneity by preparation of a periplasmic fraction, followed by ammonium sulfate fractionation and DEAE-cellulose column chromatography. Crystals were obtained by the hanging drop method using sodium citrate as a precipitant. The enzyme was found to be a dimer of identical subunits with a molecular weight of 80,000, and showed the highest activity at pH 7.5 and 45 degrees C. The enzyme was inhibited by a MAO specific inhibitor, hydroxylamine, hydrazine, phenelzine, isoniazid, and tranycpromine. The enzyme oxidized tyramine, phenethylamine, and tryptamine at higher rates, but not oxidized diamine and polyamines such as putrescine and spermine. The antibody against E. coli MAO cross-reacted with purified MAO A from Klebsiella aerogenes.
从大肠杆菌基因组文库中克隆出单胺氧化酶（MAO）基因，并使其在外周质空间中过量表达。通过制备外周质组分，随后进行硫酸铵分级分离和DEAE - 纤维素柱色谱法，将该酶纯化至同质。使用柠檬酸钠作为沉淀剂，通过悬滴法获得晶体。发现该酶是由分子量为80,000的相同亚基组成的二聚体，在pH 7.5和45℃时表现出最高活性。该酶受到MAO特异性抑制剂、羟胺、肼、苯乙肼、异烟肼和反苯环丙胺的抑制。该酶以较高速率氧化酪胺、苯乙胺和色胺，但不氧化二胺和多胺，如腐胺和精胺。抗大肠杆菌MAO的抗体与产气克雷伯菌纯化的MAO A发生交叉反应。
