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. Human Pol ? is composed of four subunits and its activity is enhanced dramatically by interaction with PCNA. The largest p125 subunit has both polymerase and 3? to 5? exonuclease activities and mutations at the p125 active sites increase genomic instability and accelerate tumorigenesis.[8,9] The remaining three subunits, p50, p68, and p12 are thought to play a regulatory role, stimulate the polymerase activity of p125 by mediating additional interactions with PCNA, and stabilize the entire Pol ? complex. p50 serves as a scaffold for the assembly of Pol ? by interacting simultaneously with all of the other three subunits. Interestingly, many essential functions of Pol ?, including the regulation of replication, trans-lesion DNA synthesis and break-induced recombination, are mediated by the third subunit. Recently we reported the crystal structure of the complex between the second p50 subunit and the 144 amino acids N-terminal domain of the third p66 subunit (p66N) of human Pol ?. However, interactions of p50/p66 with p125 and p12 subunits, as well as DNA, remain unknown. We also succeeded in prepared of soluble monodisperse samples of p125, p50/p66, and p12, and confirmed the in vitro reconstitution of four-subunit Pol ?. Our long-term goal is to reveal the structure-based molecular mechanisms of human Pol ? function. Our short-term goal is to reveal the three-dimensional organization of Pol ? complex and locate the exact position of each subunit and map the areas involved in intersubunit interactions. To reach this goal we will combine cryo EM experiments with X-ray crystallography of individual domains and confirm our conclusions with biochemical and functional experiments. The data will allow as the positioning of the recently solved X-ray structure of human p50/p66 into the electron density maps obtained by Cryo EM, thus giving more complete information about the intersubunit interactions within Pol ?. We will test the defined subunit interactions with mutations and yeast two-hybrid assay.
| Ebeida, Mohamed S; Rushdi, Ahmad A; Awad, Muhammad A et al. (2016) Disk Density Tuning of a Maximal Random Packing. Comput Graph Forum 35:259-269 |
| Wensel, Theodore G; Zhang, Zhixian; Anastassov, Ivan A et al. (2016) Structural and molecular bases of rod photoreceptor morphogenesis and disease. Prog Retin Eye Res 55:32-51 |
| Bucero, Marta Abril; Bajaj, Chandrajit; Mourrain, Bernard (2016) On the construction of general cubature formula by flat extensions. Linear Algebra Appl 502:104-125 |
| Baker, Mariah R; Fan, Guizhen; Serysheva, Irina I (2015) Single-Particle Cryo-EM of the Ryanodine Receptor Channel in an Aqueous Environment. Eur J Transl Myol 25:4803 |
| Rushdi, Ahmad A; Mitchell, Scott A; Bajaj, Chandrajit L et al. (2015) Robust All-quad Meshing of Domains with Connected Regions. Procedia Eng 124:96-108 |
| Edwards, John; Daniel, Eric; Pascucci, Valerio et al. (2015) Approximating the Generalized Voronoi Diagram of Closely Spaced Objects. Comput Graph Forum 34:299-309 |
| Wensel, Theodore G; Gilliam, Jared C (2015) Three-dimensional architecture of murine rod cilium revealed by cryo-EM. Methods Mol Biol 1271:267-92 |
| Jeter, Cameron B; Patel, Saumil S; Morris, Jeffrey S et al. (2015) Oculomotor executive function abnormalities with increased tic severity in Tourette syndrome. J Child Psychol Psychiatry 56:193-202 |
| Zhang, Qin; Cha, Deukhyun; Bajaj, Chandrajit (2015) Quality Partitioned Meshing of Multi-Material Objects. Procedia Eng 124:187-199 |
| Baker, Mariah R; Fan, Guizhen; Serysheva, Irina I (2015) Single-particle cryo-EM of the ryanodine receptor channel in an aqueous environment. Eur J Transl Myol 25:35-48 |
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