Design and Synthesis of HIV Integrase as Potential Anti-

HIV整合酶（IN）抑制剂正被开发为潜在的抗艾滋病药物。虽然大量的抑制剂已在文献中报道，很少的信息已经即将与IN蛋白的分子相互作用，这些代理商。本项目的一个重点是设计和合成药理学工具，以阐明抑制剂与IN的分子相互作用。在一项研究中，我们制备了含联苯酮的香豆素作为光亲和配体。将这些交联至IN，并通过质谱鉴定交联位点，并通过诱变和分子建模实验确认其远离IN二聚体界面处的催化位点。这些信息应有助于设计在催化位点之外起作用的界面抑制剂。另一类重要的IN抑制剂以芳基β-二酮酸为代表，其被认为在IN催化位点内起金属螯合剂的作用。为了阐明这些抑制剂与IN-DNA底物复合物相互作用的方式，将光亲和性标记物与生物素标签一起沿着在高亲和性芳基β-二酮酸抑制剂上，旨在促进光交联产物的分离和纯化。在其他研究中，HIV p6 Gag蛋白与人Tsg 101蛋白的结合已被证明是病毒出芽所必需的，并且涉及p6蛋白的关键9聚体“P-E-P-T-A-P-P-E-E”序列。我们正在准备肽和肽模拟变体的9聚体序列作为Tsg 101结合拮抗剂，可能会导致一类新的病毒出芽抑制剂。一种方法是用N-取代的甘氨酸（NSG）残基（称为“类肽”）替换Pro 4残基。然而，这是综合性的问题。因此，我们求助于一个新的类肽变体家族，其掺入了作为NSG替代物的腙酰胺。这些可以通过使一系列醛与单个HPLC纯化的酰肼前体在从固相树脂裂解后反应而容易地以文库方式进行鉴定。将这些腙还原为N-取代的“类肽酰肼”提供了一种简便的文库多样化途径。这些研究正在推进Tsg 101结合抑制剂的设计。

Inhibitors of HIV integrase (IN) are being developed as potential anti-AIDS drugs. Although a large number of inhibitors have been reported in the literature, little information has been forthcoming regarding the molecular interactions of these agents with IN protein. One focus of this project is to design and synthesize pharmacological tools to clarify molecular interactions of inhibitors with IN. In one study we prepared biphenyl ketone-containing coumarins as photoaffinity ligands. These were cross-linked to IN and the site of cross-linking was identified by mass spectroscopy and confirmed by mutagenesis and molecular modeling experiments to be distal to the catalytic site at the IN dimer interface. This information should aid in the design of interfacial inhibitors that act outside of the catalytic site. Another important class of IN inhibitors is represented by the aryl beta-diketo acids that are thought to function as metal chelators within the IN catalytic site. In order to elucidate the manner in which these inhibitors interact with IN-DNA substrate complexes, photoaffinity labels were appended onto high affinity aryl beta-diketo acid inhibitors along with biotin tags intended to facilitate isolation and purification of photo-cross-linked products. Photoactivation studies of inhibitors in the presence of IN along with MALDI-TOF mass spectral identification of cross-linked products is ongoing.In other studies, binding of the HIV p6Gag protein to human Tsg101 protein has been shown to be necessary for viral budding and to involve a critical 9-mer "P-E-P-T-A-P-P-E-E" sequence of the p6 protein. We are preparing peptide and peptide mimetic variants of this 9-mer sequence as Tsg101-binding antagonists that may lead to a new class of viral budding inhibitors. One approach was to replace the Pro4 residue with N-substitued glycine (NSG) residues (termed "peptoids"). However, this is synthetically problematic. therefore, we resorted to a new family of peptoid variants that incorporate hydrazone amides as NSG surrogates. These can be preparede readily in library fashion by reacting a series of aldehydes with a single HPLC-purified hydrazide precursor following cleavage from the solid-phase resin. Reduction of these hydrazones to N-substitued "peptoid hydrazides" affords a facile route to library diversification. These studies are advancing the design of Tsg101 binding inhibitors.