Small molecule regulation of protein conformation by binding in the Flap of HIV protease.
Small molecule regulation of protein conformation by binding in the Flap of HIV protease.
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DOI：
10.1021/cb300611p
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发表时间：
2013
影响因子：
4
通讯作者：
Stout, C. David
中科院分区：
文献类型：
--
作者：
Tiefenbrunn, Theresa;Forli, Stefano;Baksh, Michael M.;Chang, Max W.;Happer, Meaghan;Lin, Ying-Chuan;Perryman, Alexander L.;Rhee, Jin-Kyu;Torbett, Bruce E.;Olson, Arthur J.;Elder, John H.;Finn, M. G.;Stout, C. David
吲哚-6-羧酸(1F1)片段以前在基于片段的HIV蛋白酶(PR)筛选中被鉴定为翻盖结合蛋白，它已经与胃抑素抑制的PR和apo-PR共结晶。另一种片段3-吲哚丙酸(1F1-N)，由AutoDock计算预测，并在一种新的“抑制成核”结晶实验中得到证实，在两种晶体结构中利用了相同的相互作用。1F1和1F1-N都与apo-PR的闭合形式和胃抑素：PR结合。在溶液中，用差示扫描荧光法(DSF)测定1F1和1F1-N使apo-PR的Tm提高3.5-5°C，用背向散射干涉法(BSI)测定apo-PR和胃抑素：PR的微摩尔结合常数。当与apo-PR结合时，BSI中观察到的信号强度大于与胃抑素结合的PR，这与先前结合事件中更大的构象变化一致。综上所述，这些数据表明，在瓣部位的片段结合有利于HIV PR的封闭构象。
The fragment indole-6-carboxylic acid (1F1), previously identified as a flap site binder in a fragment-based screen against HIV protease (PR), has been co-crystallized with pepstatin-inhibited PR and with apo-PR. Another fragment, 3-indolepropionic acid (1F1-N), predicted by AutoDock calculations and confirmed in a novel ‘inhibition of nucleation’ crystallization assay, exploits the same interactions in the flap site in two crystal structures. Both 1F1 and 1F1-N bind to the closed form of apo-PR and to pepstatin:PR. In solution, 1F1 and 1F1-N raise the Tm of apo-PR by 3.5–5 °C as assayed by differential scanning fluorimetry (DSF), and show equivalent low-micromolar binding constants to both apo-PR and pepstatin:PR, assayed by backscattering interferometry (BSI). The observed signal intensities in BSI are greater for each fragment upon binding to apo-PR than to pepstatin-bound PR, consistent with greater conformational change in the former binding event. Together, these data indicate that fragment binding in the flap site favors a closed conformation of HIV PR.
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