Cooperative nucleic acid binding by Poly ADP-ribose polymerase 1

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Poly (ADP)-ribose polymerase 1 (PARP1) is a nuclear protein involved in DNA repair, replication, transcription, and splicing.
• PARP1's interaction with undamaged DNA and RNA is crucial for its diverse cellular functions.
• Understanding PARP1's nucleic acid binding mechanisms is key to elucidating its roles in chromatin and RNA biology.

Purpose of the Study:

• To investigate the binding characteristics of wild-type (WT) PARP1 and its mutants to various nucleic acid structures.
• To determine the role of PARP1's catalytic domain and zinc fingers in nucleic acid binding and cooperativity.
• To explore how nucleic acid secondary structure influences PARP1 binding stoichiometry and mechanism.

Main Methods:

• Analysis of PARP1 binding to single-stranded DNA (T20), single-stranded RNA (U20), and double-stranded DNA (19mer).
• Comparison of full-length WT PARP1 with mutants lacking the catalytic domain (ΔCAT) or zinc fingers 1 and 2 (ΔZn1ΔZn2).
• Characterization of protein-nucleic acid complex stoichiometry and binding patterns.

Main Results:

• WT PARP1, ΔCAT, and ΔZn1ΔZn2 PARP1 formed saturated 2:1 complexes with single-stranded oligonucleotides (T20, U20) with positive binding cooperativity.
• Neither the catalytic domain nor zinc fingers 1 and 2 are essential for cooperative binding to single-stranded nucleic acids.
• WT PARP1 formed a 4:1 complex with double-stranded DNA, while the ΔZn1Zn2 mutant showed 1:1 stoichiometry, indicating structure-dependent binding.

Conclusions:

• PARP1 exhibits structure-dependent binding mechanisms influenced by nucleic acid secondary structure.
• PARP1's interactions with diverse oligonucleotides are critical for its functional versatility in chromatin and RNA biology.
• These findings provide a foundation for understanding how PARP1 recognizes different nucleic acid ligands in cellular contexts.