Yarlett N, Garofalo J, Goldberg B, Ciminelli M A, Ruggiero V, Sufrin J R, Bacchi C J
Haskins Laboratories, Pace University, New York, NY.
Biochim Biophys Acta. 1993 Mar 24;1181(1):68-76. doi: 10.1016/0925-4439(93)90092-f.
S-adenosylmethionine synthetase was studied from bloodstream forms of Trypanosoma brucei brucei, the agent of African sleeping sickness. Two isoforms of the enzyme were evident from Eadie Hofstee and Hanes-Woolf plots of varying ATP or methionine concentrations. In the range 10-250 microM the Km for methionine was 20 microM, and this changed to 200 microM for the range 0.5-5.0 mM. In the range 10-250 microM the Km for ATP was 53 microM, and this changed to 1.75 mM for the range 0.5-5.0 mM. The trypanosome enzyme had a molecular weight of 145 kDa determined by agarose gel filtration. Methionine analogs including selenomethionine, L-2-amino-4-methoxy-cis but-3-enoic acid and ethionine acted as competitive inhibitors of methionine and as weak substrates when tested in the absence of methionine with [14C]ATP. The enzyme was not inducible in procyclic trypomastigotes in vitro, and the enzyme half-life was > 6 h. T. b. brucei AdoMet synthetase was inhibited by AdoMet (Ki 240 microM). The relative insensitivity of the trypanosome enzyme to control by product inhibition indicates it is markedly different from mammalian isoforms of the enzyme which are highly sensitive to AdoMet. Since trypanosomes treated with the ornithine decarboxylase antagonist DL-alpha-difluoromethylornithine accumulate AdoMet and dcAdoMet (final concentration approximately 5 mM), this enzyme may be the critical drug target linking inhibition of polyamine synthesis to disruption of AdoMet metabolism.
对布氏布氏锥虫（非洲昏睡病病原体）血液期的S-腺苷甲硫氨酸合成酶进行了研究。从不同ATP或甲硫氨酸浓度的伊迪·霍夫斯泰（Eadie Hofstee）和海恩斯-伍尔夫（Hanes-Woolf）图中可以明显看出该酶有两种同工型。在10 - 250微摩尔范围内，甲硫氨酸的Km为20微摩尔，在0.5 - 5.0毫摩尔范围内则变为200微摩尔。在10 - 250微摩尔范围内，ATP的Km为53微摩尔，在0.5 - 5.0毫摩尔范围内则变为1.75毫摩尔。通过琼脂糖凝胶过滤测定，锥虫酶的分子量为145 kDa。包括硒代甲硫氨酸、L - 2 - 氨基 - 4 - 甲氧基 - 顺式丁 - 3 - 烯酸和乙硫氨酸在内的甲硫氨酸类似物，在无甲硫氨酸存在的情况下用[14C]ATP进行测试时，可作为甲硫氨酸的竞争性抑制剂和弱底物。该酶在体外前循环型锥鞭毛体中不可诱导，酶的半衰期大于6小时。布氏布氏锥虫的腺苷甲硫氨酸合成酶受到腺苷甲硫氨酸的抑制（Ki为240微摩尔）。锥虫酶对产物抑制控制的相对不敏感性表明它与对腺苷甲硫氨酸高度敏感的哺乳动物同工型酶明显不同。由于用鸟氨酸脱羧酶拮抗剂DL-α-二氟甲基鸟氨酸处理的锥虫会积累腺苷甲硫氨酸和脱氧腺苷甲硫氨酸（终浓度约为5毫摩尔），这种酶可能是将多胺合成抑制与腺苷甲硫氨酸代谢破坏联系起来的关键药物靶点。
