Inhibition of Inositol Phosphorylceramide Synthase by the Cyclic Peptide Aureobasidin A
Inhibition of Inositol Phosphorylceramide Synthase by the Cyclic Peptide Aureobasidin A
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DOI：
10.1128/aac.00633-08
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发表时间：
2009-02-01
影响因子：
4.9
通讯作者：
Elhammer, Ake P.
中科院分区：
文献类型：
--
作者：
Aeed, Paul A.;Young, Casey L.;Elhammer, Ake P.
采用去污剂洗涤的膜制备物，通过对野生型和突变型酶催化反应的动力学分析，研究了真菌天然产物抑制剂aureobasidin A（阿坝）与肌醇磷酸神经酰胺合酶（IPC合酶）的相互作用。阿坝以不可逆的时间依赖性方式抑制来自白色念珠菌和酿酒酵母的野生型酶，表观Ki值分别为183和234 pM。三种合成化学衍生的阿坝衍生物PHA-533179、PHA-556655和PHA-556656的亲和力低4至5个数量级，是与供体底物磷脂酰肌醇（PI）竞争的可逆抑制剂。阿坝是一种可逆的，明显的非竞争性抑制剂，Ki为1.4 μ M，IPC合酶从阿巴抗性S。酿酒酵母突变体。当它们与突变体和野生型酶相互作用时，两种底物（神经酰胺和PI）的Km值相似。相比之下，突变酶的Vmax小于野生型酶的10%。将用阿坝获得的结果与用其他两种天然产物抑制剂（锈菌菌素和卡氟芬净）获得的结果进行比较，发现虽然锈菌菌素似乎是野生型酶的可逆的、非竞争性抑制剂，Ki为16.0 nM，但卡氟芬净具有时间依赖性抑制剂的动力学性质，表观Ki为0.43 nM。这些化合物作为突变酶抑制剂的效率的评价显示，对于两者，对酶的表观亲和力下降超过2个数量级。
By using a detergent-washed membrane preparation, the interaction of the fungal natural product inhibitor aureobasidin A (AbA) with inositol phosphorylceramide synthase (IPC synthase) was studied by kinetic analysis of wild-type and mutant enzyme-catalyzed reactions. AbA inhibited the wild-type enzyme from both Candida albicans and Saccharomyces cerevisiae in an irreversible, time-dependent manner, with apparent K-i values of 183 and 234 pM, respectively. Three synthetic chemistry-derived AbA derivatives, PHA-533179, PHA-556655, and PHA-556656, had affinities 4 to 5 orders of magnitude lower and were reversible inhibitors that competed with the donor substrate phosphatidylinositol (PI). AbA was a reversible, apparently noncompetitive inhibitor, with a Ki of 1.4 mu M, of the IPC synthase from an AbA-resistant S. cerevisiae mutant. The K-m values for both substrates (ceramide and PI) were similar when they interacted with the mutant and the wild-type enzymes. By contrast, the V-max for the mutant enzyme was less than 10% of that for the wild-type enzyme. A comparison of the results obtained with AbA with those obtained with two other natural products inhibitors, rustmicin and khafrefungin, revealed that while rustmicin appeared to be a reversible, noncompetitive inhibitor of the wild-type enzyme, with a K-i of 16.0 nM, khafrefungin had the kinetic properties of a time-dependent inhibitor and an apparent K-i of 0.43 nM. An evaluation of the efficiencies of these compounds as inhibitors of the mutant enzyme revealed for both a drop in the apparent affinity for the enzyme of more than 2 orders of magnitude.