Thermodynamic dissection of the binding energetics of KNI-272, a potent HIV-1 protease inhibitor
Thermodynamic dissection of the binding energetics of KNI-272, a potent HIV-1 protease inhibitor
复制标题
DOI：
10.1110/ps.9.9.1801
复制
发表时间：
2000-09-01
期刊：
影响因子：
8
通讯作者：
Freire, E
中科院分区：
文献类型：
--
作者：
Velazquez-Campoy, A;Luque, I;Freire, E
KNI-272是一种强效的HIV-1蛋白酶抑制剂，据报道其抑制常数在皮摩尔范围内。在本文中，一个完整的实验解剖的热力学力量，定义这种抑制剂的结合亲和力的野生型和耐药突变体V82 F/I84 V。与其他蛋白酶抑制剂不同，KNI-272以有利的结合焓与蛋白酶结合。有利的结合焓的起源已被追溯到耦合的结合反应的埋葬的六个水分子。这些结合水分子，以前确定的NMR研究，优化原子包装在抑制剂/蛋白质界面增强货车德瓦尔斯和其他有利的相互作用。这些相互作用抵消了通常与疏水分子结合相关的不利焓。与耐药突变体的缔合常数弱100-500倍。结合亲和力的降低对应于结合吉布斯能增加3-3.5 kcal/mol，这源于较不利的焓变（1.7 kcal/mol更正）和熵变。量热结合实验作为pH值的函数进行，并利用不同的离子化缓冲液，允许解剖的质子连接的影响。根据这些实验，抑制剂的结合与两个基团的质子化/去质子化有关。在未复合形式中，这些基团的pK为6.0和4.8，而在复合物中变为6.6和2.9。这些基团已被鉴定为蛋白酶中催化的异丙基二联体和抑制剂分子中的异喹啉氮中的一种异丙基酯。结合亲和力在pH 5和pH 6之间最大。在这些pH值下，亲和力接近6 × 10（10）M-1（K-d = 16 pM）。数据的总体分析得出缓冲液和pH无关的结合焓为-6.3 kcal/mol。在质子交换为零的条件下，结合的吉布斯能为-14.7千卡/摩尔，由此得到结合熵为28千卡/K摩尔。因此，KNI-272的结合在热力学上和熵上都是有利的。基于结构的热力学分析表明，KNI-272的别苯基-去甲他汀核为设计对耐药突变不敏感的抑制剂提供了重要的支架。
KNI-272 is a powerful HTV-1 protease inhibitor with a reported inhibition constant in the picomolar range. In this paper, a complete experimental dissection of the thermodynamic forces that define the binding affinity of this inhibitor to the wild-type and drug-resistant mutant V82F/I84V is presented. Unlike other protease inhibitors, KNI-272 binds to the protease with a favorable binding enthalpy. The origin of the favorable binding enthalpy has been traced to the coupling of the binding reaction to the burial of six water molecules. These bound water molecules, previously identified by NMR studies, optimize the atomic packing at the inhibitor/protein interface enhancing van der Waals and other favorable interactions. These interactions offset the unfavorable enthalpy usually associated with the binding of hydrophobic molecules. The association constant to the drug resistant mutant is 100-500 times weaker. The decrease in binding affinity corresponds to an increase in the Gibbs energy of binding of 3-3.5 kcal/mol, which originates from less favorable enthalpy (1.7 kcal/mol more positive) and entropy changes. Calorimetric binding experiments performed as a function of pH and utilizing buffers with different ionization enthalpies have permitted the dissection of proton linkage effects. According to these experiments, the binding of the inhibitor is linked to the protonation/deprotonation of two groups. in the uncomplexed form these groups have pKs of 6.0 and 4.8, and become 6.6 and 2.9 in the complex. These groups have been identified as one of the aspartates in the catalytic aspartyl dyad in the protease and the isoquinoline nitrogen in the inhibitor molecule. The binding affinity is maximal between pH 5 and pH 6. At those pH values the affinity is close to 6 x 10(10) M-1 (K-d = 16 pM) Global analysis of the data yield a buffer- and pH-independent binding enthalpy of -6.3 kcal/mol. Under conditions in which the exchange of protons is zero, the Gibbs energy of binding is -14.7 kcal/mol from which a binding entropy of 28 cal/K mol is obtained. Thus, the binding of KNI-272 is both enthalpically and entropically favorable. The structure-based thermodynamic analysis indicates that the allophenyl-norstatine nucleus of KNI-272 provides an important scaffold for the design of inhibitors that are less susceptible to resistant mutations.