Subtype Polymorphisms Among HIV-1 Protease Variants Confer Altered Flap Conformations and Flexibility
Subtype Polymorphisms Among HIV-1 Protease Variants Confer Altered Flap Conformations and Flexibility
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DOI：
10.1021/ja907088a
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发表时间：
2009-10-21
影响因子：
15
通讯作者：
Fanucci, Gail E.
中科院分区：
文献类型：
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作者：
Kear, Jamie L.;Blackburn, Mandy E.;Fanucci, Gail E.
人类免疫缺陷病毒 1 型 (HIV-1) 蛋白酶在逆转录病毒 HIV-1 的成熟和生命周期中发挥着重要作用，因为它在调节病毒多蛋白 gag 和 gag-pol 的翻译后加工中发挥作用；因此，它是艾滋病抗病毒治疗的一个关键目标。底物与活性位点的可及性由两个瓣介导，这两个瓣必须经历从开放构象到闭合构象的大构象变化（引诱底物结合和催化）。利用电子顺磁共振（EPR）方法和双电子-电子共振（DEER）光谱的定点自旋标记（SDSL）来监测apo HIV-1中瓣的构象 蛋白酶 (HIV-1PR)、B、C 和 F 亚型、CRF01_A/E 以及患者分离株 V6 和 MDR 769。重建从偶极调制回波曲线分析中获得的距离分布图，以产生一组高斯形群体，从而对采样的皮瓣构象进行分析。每个构象异构体群体的相对百分比被描述为“折叠/弯曲”， 确定并比较各种构建体的“封闭”、“半开放”和“全开放”。结果和分析表明，apo HIV-1PR 亚型、CRF 和患者分离株之间的序列变异改变了平均瓣构象，这种方式可以理解为诱导先前描述的 HIV-1PR 四种构象的相对群体或构象采样的变化。
Human immunodeficiency virus type 1 (HIV-1) protease plays a fundamental role in the maturation and life cycle of the retrovirus HIV-1, as it functions in regulating post-translational processing of the viral polyproteins gag and gag-pol; thus, it is a key target of AIDS antiviral therapy. Accessibility of substrate to the active site is mediated by two flaps, which must undergo a large conformational change from an open to a closed conformation (luring substrate binding and catalysis. The electron paramagnetic resonance (EPR) method of site-directed spin labeling (SDSL) with double electron-electron resonance (DEER) spectroscopy was utilized to monitor the conformations of the flaps in apo HIV-1 protease (HIV-1PR), subtypes B, C, and F, CRF01_A/E, and patient isolates V6 and MDR 769. The distance distribution profiles obtained from analysis of the dipolar modulated echo curves were reconstructed to yield a set of Gaussian-shaped populations, which provide an analysis of the flap conformations sampled. The relative percentages of each conformer population described as "tucked/curted", "closed", "semi-open", and "wide-open" were determined and compared for various constructs. The results and analyses show that sequence variations among subtypes, CRFs, and patient isolates of apo HIV-1PR alter the average flap conformation in a way that can be understood as inducing shifts in the relative populations, or conformational sampling, of the previously described four conformations for HIV-1PR.