Structural Studies of SUMO Protein Modification

描述(由申请人提供)：用小的泛素样修饰物SUMO修饰细胞蛋白质对酵母中真核细胞的核过程和细胞周期进程是必不可少的。与泛素结合不同，泛素结合通常针对蛋白质被蛋白酶体降解，相扑修饰针对蛋白质活性和定位的变化。相扑结合至少由三种酶催化，E1是相扑激活酶，E2是相扑结合酶，ULPS是相扑蛋白水解酶和去共轭处理酶。SUMO被翻译为前体，需要C-末端的ULP蛋白降解才能产生成熟的形式。SUMO通过C-末端的腺基化被E1激活，然后转移到分子内的半胱氨酸，在SUMO和E1之间形成硫酯键。然后，将E1-相扑硫酸酯转移到E2半胱氨酸上，在E2和相扑之间形成硫酯键。E2-相扑复合体能够将相扑转移到蛋白质靶标上的赖氨酸受体上，形成稳定的异肽链。相扑修饰是可逆的，它从赖氨酸中去除是由U1p酶催化的。相扑结合的调控途径包括激活哺乳动物RanGAP1、P53转录调控、MDM2泛素连接酶调控、I-kappa-B-α泛素化保护、PML、PML-RAR和SP100核定位、着丝粒分离和间隔素环形成，表明相扑结合是参与核代谢和细胞周期控制的细胞过程中的中心调节因子。由于相扑结合途径的分子细节知之甚少，因此我们将利用结构、生化和遗传技术来建立相扑过程的物理模型，通过表征相扑、E2介导的相扑、E2-靶蛋白复合体以及相扑结合蛋白、结合酶和脱偶联酶之间的复合体。通过鉴定新的相扑修饰蛋白，将建立相扑甲基化的遗传和生化模型。对相扑途径的彻底结构和功能分析将使人们对其潜在的生物学有更深入的了解。

DESCRIPTION (provided by applicant): Modification of cellular proteins by the small Ubiquitin-like Modifier SUMO is essential for eukaryotic nuclear processes and cell cycle progression in yeast. Unlike ubiquitin conjugation, which generally targets proteins for degradation by the proteasome, SUMO modification targets proteins for changes in activity and localization. SUMO conjugation is catalyzed by at least three enzymes, E1, the SUMO activating enzyme, E2, the SUMO conjugation enzyme, and Ulps, SUMO proteolytic processing and deconjugation enzymes. SUMO is translated as a precursor, requiring C-terminal Ulp proteolysis to generate the mature form. SUMO is activated via C-terminal adenylation by E1 and subsequently transferred to an intramolecular cysteine, forming a thioester bond between SUMO and E1. The E1-SUMO thioester is then transferred to an E2 cysteine forming a thioester bond between E2 and SUMO. The E2-SUMO complex is competent to transfer SUMO to lysine receptors on the protein target, forming a stable isopeptide linkage. SUMO modification is reversible, and its removal from lysine is catalyzed by U1p proteases. Pathways modulated by SUMO conjugation include activation of mammalian RanGAP1, p53 transcriptional regulation, MDM2 ubiquitin ligase regulation, I-kappa-B-alpha protection from ubiquitination, PML, PML-RAR, and SP100 nuclear localization, centromere segregation, and septin ring formation, indicating SUMO conjugation as a central regulator in cellular processes involved in nuclear metabolism and cell cycle control.Since few molecular details are known about the SUMO conjugation pathway, structural, biochemical and genetic techniques will be utilized to establish physical models for the sumoylation process by characterizing E1 activation of SUMO, E2 mediated SUMO conjugation, E2-target protein complexes, and complexes between sumoylated proteins, conjugating, and deconjugating enzymes. Genetic and biochemical models will be established for sumoylation through identification of novel SUMO modified proteins. A thorough structure-function analysis of the SUMO pathway will yield insights into its underlying biology.