HUMAN PYRUVATE KINASE ISOFORM 2 BINDING

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 丙酮酸激酶是一种重要的糖酵解酶，其活性受到高度调控。该酶有4种亚型。R亚型和L亚型分别针对红细胞和肝脏。另外两种异构体M1和M2是同一基因的剪接变体，只有一个外显子不同。虽然M2亚型在胚胎细胞和成人干细胞中高度表达，但大多数成人组织表达M1亚型。然而，所有的癌细胞都被发现完全恢复了M2的表达。尽管这种转变的原因目前尚不清楚，但正常组织和肿瘤之间的这种差异表达使M2亚型成为癌症治疗的药物靶点。我们已经发现了几种化合物，它们特异性地抑制M2亚型，而不是M1亚型。由于先导化合物仍具有相当高的IC50，我们希望确定与缓蚀剂结合的M2的结构，以便进行合理的修饰以产生更好的化合物。此外，与PKM1或L亚型不同，PKM2优先与磷酸酪氨酸肽库相互作用。从诱变研究中，我们还发现这种相互作用降低了它的酶活性，可能是通过取代变构激活剂FBP。这可能解释了为什么癌细胞会有PKM2亚型，因为额外的控制水平有利于癌细胞对营养物质的剥夺做出反应。为了更好地了解PKM2活性的调节，我们希望在原子水平上看到磷酸肽是如何与FBP竞争的。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Pyruvate kinase is an essential glycolytic enzyme whose activity is highly regulated. There are 4 isoforms of the enzyme. The R and L isoforms are specific for red blood cells and liver respectively. The other two isoforms, M1 and M2, are splice variant of the same gene with only one exon difference. While the M2 isoform is highly expressed in embryonic cells and adult stem cells, most adult tissues express the M1 isoform. However, all cancer cells are found to switch back completely M2 expression. Although the cause of this switch is currently unknown, this differential expression between normal tissues and tumor makes the M2 isoform a drug target for cancer treatment. We have found several compounds that specifically inhibit the M2 isoform and not the M1 isoform. Since the lead compounds still have fairly high IC50, we would like to determine the structure of M2 bound to the inhibitors so rational modification can be made to generate better compounds. In addition, in contrast to PKM1 or the L isoforms, PKM2 preferentially interacts with phosphotyrosine peptide libraries. From mutagenesis studies, we also found that this interaction reduces its enzyme activity, presumably by displacing the allosteric activator FBP. This may explain why cancer cells have PKM2 isoform, since the extra level of control is advantageous for cancer cells to respond to deprivation of nutrients. In order to understand the regulation of PKM2 activity better, we would like to see at the atomic level, how the phosphopeptide competes with FBP.