Dataset Information

Structural and biophysical studies of human PARP-1 in complex with damaged DNA.

ABSTRACT: The enzyme poly(ADP-ribose) polymerase-1 (PARP-1) is a global monitor of chromatin structure and DNA damage repair. PARP-1 binds to nucleosomes and poly(ADP-ribosylates) histones and several chromatin-associated factors to expose specific DNA sequences to the cellular machinery involved in gene transcription and/or DNA damage repair. While these processes are critical to genomic stability, the molecular mechanisms of how DNA damage induces PARP-1 activation are poorly understood. We have used biochemical and thermodynamic measurements in conjunction with small-angle X-ray scattering to determine the stoichiometry, affinity, and overall structure of a human PARP-1 construct containing the entire DNA binding region, the zinc ribbon domain, and automodification domains (residues 1-486). The interaction of this PARP-1 protein construct with three different DNA damage models (DNA constructs containing a nick, a blunt end, or a 3' extension) was evaluated. Our data indicate that PARP-1 binds each DNA damage model as a monomer and with similar affinity, in all cases resulting in robust activation of the catalytic domain. Using small-angle X-ray scattering, we determined that the N-terminal half of PARP-1 behaves as an extended and flexible arrangement of individually folded domains in the absence of DNA. Upon binding DNA, PARP-1 undergoes a conformational change in the area surrounding the zinc ribbon domain. These data support a model in which PARP-1, upon binding DNA, undergoes a conformational change to become an active nuclear enzyme.

SUBMITTER: Lilyestrom W

PROVIDER: S-EPMC2864582 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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Structural and biophysical studies of human PARP-1 in complex with damaged DNA.

Lilyestrom Wayne W van der Woerd Mark J MJ Clark Nicholas N Luger Karolin K

Journal of molecular biology 20091204 5

The enzyme poly(ADP-ribose) polymerase-1 (PARP-1) is a global monitor of chromatin structure and DNA damage repair. PARP-1 binds to nucleosomes and poly(ADP-ribosylates) histones and several chromatin-associated factors to expose specific DNA sequences to the cellular machinery involved in gene transcription and/or DNA damage repair. While these processes are critical to genomic stability, the molecular mechanisms of how DNA damage induces PARP-1 activation are poorly understood. We have used bi ...[more]

PMID: 19962992

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Project description:A pair of stereoisomeric covalent adducts to guanine in double-stranded DNA, derived from the reaction of mutagenic and tumorigenic metabolites of benzo[a]pyrene, have been well characterized structurally and thermodynamically. Both high-resolution NMR solution structures and an array of thermodynamic data are available for these 10S (+)- and 10R (-)-trans-anti -[BP]-N(2)-dG adducts in double-stranded deoxyoligonucleotides. The availability of experimentally well-characterized duplexes containing these two stereoisomeric guanine adducts provides an opportunity for evaluating the molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) method for computing thermodynamic properties from molecular dynamics ensembles. We have carried out 3-ns molecular dynamics simulations, using NMR solution structures as the starting models for the 10S (+)- and 10R (-)-trans-anti-dG adducts in a DNA duplex 11-mer using AMBER 6.0. We employed the MM-PBSA method to compute the free energies, enthalpies, and entropies of the two adducts. Our complete thermodynamic analysis agrees quite well with the full experimental thermodynamic characterization of these adducts, showing essentially equal stabilities of the two adducts. We also calculated the nuclear Overhauser effect (NOE) distances from the molecular dynamics trajectories, and compared them against the experimental NMR-derived NOE distances. Our results showed that the simulated structures are in good agreement with the NMR experimental NOE data. Furthermore, the molecular dynamics simulations provided new structural and biological insights. Specifically, the puzzling observation that the BP aromatic ring system in the 10S (+)-trans-anti-dG adduct is more exposed to the aqueous solvent than the 10R (-)-trans-anti-dG adduct, is rationalized in terms of the adduct structures. The structural and thermodynamic features of these stereoisomeric adducts are also discussed in relation to their reported low susceptibilities to nucleotide excision repair.

Dataset Information

Structural and biophysical studies of human PARP-1 in complex with damaged DNA.

Publications

Structural and biophysical studies of human PARP-1 in complex with damaged DNA.

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