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Moor, N. A.

Publications and source records attributed to Moor, N. A..

3 recordsLinked to original sources

More than just a passive brick in the wall: the nucleosome facilitates DNA polymerase β activity in linker DNA and its PARP-dependent regulation in the BER pathway choice

DNA polymerase {beta} (Pol{beta}) is a central player of base excision repair (BER), performing gap-filling synthesis on damaged DNA. While nucleosome core particles (NCPs) are known to impede activity of BER enzymes, the regulation of this process in linker DNA adjacent to nucleosomes remains unclear. Here we demonstrate an unexpected stimulation of Pol{beta}-catalyzed gap-filling and strand-displacement synthesis in linker DNA by the adjacent NCP. Notably, the nucleosomal context reinforces the regulatory modulation of Pol{beta} activity by PARP1/PARP2 and FEN1. While linker histone H1 restricts strand-displacement synthesis at the nucleosome entry/exit site, PARP1 and PARP2 modulate Pol{beta} function through competitive binding to DNA gaps or nicks and via poly(ADP-ribosyl)ation (PARylation). At the same time, PARPs binding differentially regulates BER sub-pathway choice, and PARylation alleviates H1-mediated inhibition. These findings reveal a multi-layered regulatory system wherein the nucleosome acts as a dynamic platform coordinating Pol{beta} activity and its interplay with chromatin-associated factors, influencing the balance between short- and long-patch BER. The research advances understanding of chromatin-mediated control of BER DNA repair synthesis and the functional specialization of PARP1 and PARP2 in maintaining genome stability. HighlightsThe nucleosome core particle acts not only as a barrier but also as a stimulator of Pol{beta}-mediated DNA repair synthesis in adjacent linker DNA. The nucleosome acts as an allosteric platform that enhances the regulatory functions of chromatin-associated factors (PARP1, PARP2, H1) in the linker DNA repair synthesis. PARP1 suppresses overall Pol{beta} synthesis, while PARP2 specifically inhibits strand displacement, thereby gating the choice between short- and long-patch BER pathways.

molecular biology↗

Cas9 is mostly orthogonal to human systems of DNA break sensing and repair

CRISPR/Cas9 system is [a] powerful gene editing tool based on the RNA-guided cleavage of target DNA. The Cas9 activity can be modulated by proteins involved in DNA damage signalling and repair due to their interaction with double- and single-strand breaks (DSB and SSB, respectively) generated by wild-type Cas9 or Cas9 nickases. Here we address the interplay between Streptococcus pyogenes Cas9 and key DNA repair factors, including poly(ADP-ribose) polymerase 1 (SSB/DSB sensor), its closest homolog poly(ADP-ribose) polymerase 2, Ku antigen (DSB sensor), DNA ligase I (SSB sensor), replication protein A (DNA duplex destabilizer), and Y-box binding protein 1 (RNA/DNA binding protein). None of those significantly affected Cas9 activity, while Cas9 efficiently shielded DSBs and SSBs from their sensors. Poly(ADP-ribosyl)ation of Cas9 detected for poly(ADP-ribose) polymerase 2 had no apparent effect on the activity. In cellulo, Cas9-dependent gene editing was independent of poly(ADP-ribose) polymerase 1. Thus, Cas9 can be regarded as an enzyme mostly orthogonal to the natural regulation of human systems of DNA break sensing and repair.

biochemistry↗

Structural insights into tick-borne encephalitis virus neutralization and animal protection by a therapeutic antibody

Tick-borne encephalitis virus (TBEV) causes about 5-6 thousand cases annually, while there is still no effective treatment for this virus. To fill this gap, a high-affinity chimeric anti-TBEV antibody ch14D5 has previously been constructed, and high protective activity in a murine TBEV model has been shown for this antibody. However, the mechanism of action of this antibody and the recognized epitope have not been known yet. In this study, it is shown by X-ray crystallography that this antibody recognizes a unique epitope on the lateral ridge of the D3 domain of glycoprotein E, which is readily accessible for binding. The orientation of this antibody relative to the virion surface makes bivalent binding possible, which facilitates the cross-linking of glycoprotein E molecules and thus blocking of surface rearrangements required for infection. Since the antibody tightly binds to this protein even at pH [~] 5.0, it locks the virion in an acidic environment inside the late endosomes or phagosomes and, therefore, effectively blocks the fusion of the viral and endosomal/phagosomal membranes. We believe that this is why the ch14D5 antibody does not induce an antibody-dependent enhancement of infection in vivo, which is critical in the development of antibody-based therapeutic agents. In addition, the structure of the antibody-glycoprotein E interface can be used for the rational design of this antibody for enhancing its properties.

biochemistry↗