Template-competitive inhibitors of HIV-1 reverse transcriptase: design, synthesis and inhibitory activity.
Template-competitive inhibitors of HIV-1 reverse transcriptase: design, synthesis and inhibitory activity.
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HIV-1 逆转录酶的模板竞争性抑制剂：设计、合成和抑制活性。
DOI：
10.1016/s0968-0896(01)00297-8
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发表时间：
2002
影响因子：
3.5
通讯作者：
Doughty,MichaelB
中科院分区：
文献类型：
--
作者：
Li,Ke;Lin,Weiying;Chong,KarHua;Moore,BobM;Doughty,MichaelB
我们报道了一类新型模板竞争性逆转录酶抑制剂（TCRTIs）的设计、合成和活性研究。TCRTI是在我们的实验室中合成的一系列基于dATP的模板竞争性DNA聚合酶抑制剂的1，N6-乙烯基类似物（摩尔，B. M.; Jenyi，R.;道蒂，M.B. Biochemistry 1996，35，11634）。因此，核苷酸2-（4-叠氮苯甲酰基）硫代-1，N6-亚乙基-2 ′-脱氧腺苷5′-三磷酸1，四氟类似物2-以2-硫代-1，3，5，6-四氟苯甲酰基-4-叠氮基）硫代-1，N6-亚乙基-2 ′-脱氧腺苷5′-三磷酸2为原料，N6-亚乙基-2 ′-脱氧腺苷5′-单磷酸与相应的氯代或溴代烷基卤反应，转化为三磷酸。在动力学上，核苷酸1和2都是逆转录酶对模板/引物的竞争性抑制剂，Ki分别为8.0和7.4μM，而对TTP的非竞争性抑制剂，Ki分别为15和10μM。核苷酸3，它不同于1只在它缺乏etheno组，非互补核苷酸三磷酸，和相关的单磷酸和核苷，是完全无活性的逆转录酶抑制剂的浓度高达1 mM。光灭活RT由1是时间和浓度依赖性的，并保护模板/引物，但不是由dNTPs。浓度依赖性失活数据显示KD约为17.2 μM，最大失活率为90%，放射性标记的[β，γ-32 P]-1特异性共价结合到RT的p66亚基中。因此，光失活数据支持我们从动力学数据得出的主要结论，即这类RT抑制剂是非底物和模板竞争性的。
We report the design, synthesis and activity studies on a novel class of template-competitive reverse transcriptase inhibitors (TCRTIs). The TCRTIs are 1,N6-etheno analogues of a series of dATP-based template-competitive DNA polymerase inhibitors synthesized in our laboratory (Moore, B. M.; Jalluri, R.; Doughty, M.B. Biochemistry1996, 35, 11634). Thus, nucleotides 2-(4-azidophenacyl)thio-1,N6-etheno-2′-deoxyadenosine 5′-triphosphate 1, the tetrafluoro analogue 2-(4-azido-2,3,5,6-tetrafluorophenacyl)thio-1,N6-etheno-2′-deoxyadenosine 5′-triphosphate 2 and its analogues were synthesized by alkylation of 2-thio-1,N6-etheno-2′-deoxyadenosine 5′-monophosphate with the corresponding chloro- or bromo-alkyl halides and converted to the triphosphate. Kinetically, nucleotides 1 and 2 are both competitive inhibitors of reverse transcriptase versus template/primer with Ki's of 8.0 and 7.4μM, respectively, and non-competitive inhibitors versus TTP with Ki's of 15 and 10μM, respectively. Nucleotide 3, which differs from 1 only in that it lacks the etheno group, non-complementary nucleotide triphosphates, and related monophosphates and nucleosides, are completely inactive as inhibitors of reverse transcriptase at concentrations up to 1 mM. Photoinactivation of RT by 1 was both time- and concentration-dependent, and protected by template/primer but not by dNTPs. The concentration-dependent inactivation data gave a KD,appof 17.2 μM and maximum inactivation of 90%, and radiolabeled [β,γ-32P]-1 photoincorporated specifically and covalently into the p66 subunit of RT. Thus the photoinactivation data support our main conclusion from the kinetic data that this class of RT inhibitors are non-substrate and template-competitive.