Structural Studies of SUMO Protein Modification

描述（由申请人提供）：小泛素样修饰物SUMO对细胞蛋白质的修饰对于酵母中的真核核细胞核过程和细胞周期进程至关重要。与泛素缀合不同，泛素缀合通常靶向蛋白质以通过蛋白酶体降解，SUMO修饰靶向蛋白质以改变活性和定位。SUMO缀合由至少三种酶催化，E1，SUMO活化酶，E2，SUMO缀合酶，和Ulps，SUMO蛋白水解加工和解缀合酶。SUMO被翻译为前体，需要C-末端Ulp蛋白水解以产生成熟形式。SUMO通过C-末端腺苷酸化被E1激活，随后转移到分子内半胱氨酸，在SUMO和E1之间形成硫酯键。然后将E1-SUMO硫酯转移至E2半胱氨酸，在E2和SUMO之间形成硫酯键。E2-SUMO复合物能够将SUMO转移到蛋白质靶点上的赖氨酸受体，形成稳定的异肽连接。SUMO修饰是可逆的，其从赖氨酸上的去除由U1 p蛋白酶催化。通过SUMO缀合调节的途径包括哺乳动物RanGAP 1的激活、p53转录调节、MDM 2泛素连接酶调节、I-κ-B-α对泛素化的保护、PML、PML-RAR和SP100核定位、着丝粒分离和隔蛋白环形成，这表明SUMO结合作为参与核代谢和细胞周期控制的细胞过程的中心调节器。由于已知SUMO结合途径，将利用结构、生物化学和遗传技术通过表征SUMO的E1活化、E2介导的SUMO结合、E2-靶蛋白复合物以及SUMO化蛋白、结合和去结合酶之间的复合物来建立SUMO化过程的物理模型。通过鉴定新的SUMO修饰蛋白，将建立SUMO化的遗传和生化模型。对SUMO通路进行彻底的结构-功能分析将有助于深入了解其潜在的生物学。

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.