DNA-Dependent Protein Kinase in Non-Homologous End-Joining: Guarding Strategic Positions

__Abstract__ Careful maintenance of genetic information throughout generations is of vital importance to all living creatures. A battery of both endogenous and exogenous factors continuously threatens genetic integrity by altering the DNA chemistry. As a consequence, DNA damage types are as diverse as their causes. DNA doublestrand breaks (DSBs) are among the most deleterious lesions, since they introduce chromosomal breakage or translocation and are able to trigger carcinogenesis. Perhaps even more importantly, DSBs may cause either cell death or permanent growth arrest. Fortunately, the mammalian cell has two effective DSB repair mechanisms at its disposal: homologous recombination (HR) and non-homologous end-joining (NHEJ). The research efforts presented in this thesis contribute to the understanding of the molecular mechanism of NHEJ in general, and the function of one of its core enzyme complexes, DNA-dependent protein kinase (DNA-PK), in particular

DNA-binding polarity of human replication protein A positions nucleases in nucleotide excision repair

The human single-stranded DNA-binding replication A protein (RPA) is involved in various DNA-processing events. By comparing the affinity of hRPA for artificial DNA hairpin structures with 3'- or 5'-protruding single-stranded arms, we found that hRPA binds ssDNA with a defined polarity; a strong ssDNA interaction domain of hRPA is positioned at the 5' side of its binding region, a weak ssDNA-binding domain resides at the 3' side. Polarity appears crucial for positioning o