Cell-free assay technologies for the identification of active compounds

PCMBS与MTP检测开发和筛选部门密切合作 (ADSS/Henrich）自动化、验证和完成高通量筛选（HTS）， 化学品多样性和发展科（CDDS/Beutler）获得新的化学品来源 多样性，并与天然产品化学科（NPCS/Grkovic，Du）优先考虑 用于天然产物分离和纯活性化合物生物化学评价的提取物 通过NPCS或CDDS分离。在此审查期间，PCMBS工作的示例包括 发表的关于筛选蛋白酶MALT的选择性抑制剂的天然产物的论文 泛素连接酶CBLB和磷酸二酯酶TDP 1。我们对天然产物的研究 TDP 1抑制剂还导致了第二篇论文描述了一类新的环肽 它们是该磷酸二酯酶的第一个报道的变构抑制剂。我们最近 也完成了磷酸二酯酶TDP 2抑制剂的筛选， 准备.此外，住房和抵押贷款委员会继续加强执行 用于鉴定热传导的特定调节剂的生物物理筛选方法 蛋白质和RNA结构的稳定性。最初，这项工作导致了 鉴定一种新的小分子支架，改变HIV-1 TAR RNA的稳定性。 这项技术现在已经扩展到可以方便地筛选纯化合物 和天然产物样品以高通量形式鉴定RNA调节剂， 蛋白质稳定性最近发表的一个例子是鉴定一种天然产物 选择性调节pre-miR-21的热稳定性。我们目前有两个新的 针对嵌合激酶DNAJB 1-PRKACA（PKADJ）和肌醇的酶筛选正在进行中 1，3，4-三磷酸5/6激酶（ITPK 1），以及三个新的生物物理筛选;针对 免疫相关代谢酶免疫应答基因1（IRG 1），诊断 肝细胞癌磷脂酰肌醇蛋白聚糖-3（GPC-3）与SARS-CoV-2受体结合的靶点 域（RBD）。PCMBS还评估了来自MTP筛选的化合物的特异性 与大分子靶点的相互作用。我们使用了生物物理和生物化学的方法 评价pre-miR-21 RNA 6、TDP 1，3，9 MALT 1，5和SARS-CoV-2病毒抑制剂的技术 入境

The PCMBS collaborates closely with the MTP Assay Development and Screening Section (ADSS/Henrich) to automate, validate and complete high throughput screens (HTS), with the Chemical Diversity and Development Section (CDDS/Beutler) to access new sources of chemical diversity and with the Natural Products Chemistry Section (NPCS/Grkovic, Du) to prioritize extracts for natural product isolation and biochemically evaluate pure active compounds isolated by NPCS or CDDS. Examples of PCMBS efforts during this review period, include published papers on screens of natural products for selective inhibitors of the protease MALT the ubiquitin ligase CBLB, and the phosphodiesterase TDP1. Our studies on natural product inhibitors of TDP1 also resulted in a second paper describing a new class of cyclic peptides which are the first reported allosteric inhibitors of this phosphodiesterase. We have recently also completed a screen for inhibitors of the phosphodiesterase TDP2 with a manuscript in preparation. In addition, the PCMBS has continued to increase the implementation of biophysical screening methods for the identification of specific modulators of the thermal stability of both protein and RNA structures. Initially, this work resulted in the identification of a new small molecule scaffold that altered the stability of HIV-1 TAR RNA. This technology has now been expanded upon to allow for the facile screening of pure compounds and natural product samples in a high throughput format to identify modulators of RNA and protein stability. One recently published example was the identification of a natural product that selectively modulated the thermal stability of pre-miR-21. We currently have two new enzymatic screens in progress, against the chimeric kinase DNAJB1-PRKACA (PKADJ) and inositol 1,3,4-triphosphate 5/6 kinase (ITPK1), as well as three new biophysical screens; against the immunologically-pertinent metabolic enzyme Immune Responsive Gene 1 (IRG1), the diagnostic target for hepatocellular carcinoma glypican-3 (GPC-3) and the SARS-CoV-2 receptor binding domain (RBD). The PCMBS also evaluates compounds derived from MTP screens for their specific interactions against macromolecular targets. We have used both biophysical and biochemical techniques to evaluate inhibitors of pre-miR-21 RNA6, TDP1,3,9 MALT1,5 and SARS-CoV-2 viral entry.