Mechanisms Of protein-DNA Interactions Research Articles

EGPDI: identifying protein-DNA binding sites based on multi-view graph embedding fusion.

Mechanisms of protein-DNA interactions are involved in a wide range of biological activities and processes. Accurately identifying binding sites between proteins and DNA is crucial for analyzing genetic material, exploring protein functions, and designing novel drugs. In recent years, several computational methods have been proposed as alternatives to time-consuming and expensive traditional experiments. However, accurately predicting protein-DNA binding sites still remains a challenge. Existing computational methods often rely on handcrafted features and a single-model architecture, leaving room for improvement. We propose a novel computational method, called EGPDI, based on multi-view graph embedding fusion. This approach involves the integration of Equivariant Graph Neural Networks (EGNN) and Graph Convolutional Networks II (GCNII), independently configured to profoundly mine the global and local node embedding representations. An advanced gated multi-head attention mechanism is subsequently employed to capture the attention weights of the dual embedding representations, thereby facilitating the integration of node features. Besides, extra node features from protein language models are introduced to provide more structural information. To our knowledge, this is the first time that multi-view graph embedding fusion has been applied to the task of protein-DNA binding site prediction. The results of five-fold cross-validation and independent testing demonstrate that EGPDI outperforms state-of-the-art methods. Further comparative experiments and case studies also verify the superiority and generalization ability of EGPDI.

NMR Studies of Restriction Enzyme−DNA Interactions: Role of Conformation in Sequence Specificity

Sequence specific DNA binding proteins are thought to adopt distinct conformations when binding to target (cognate) and nontarget (noncognate) sequences. There is both biochemical and crystallographic evidence that this behavior is important in mediating sequence recognition by the Mg(II)-dependent type II restriction enzymes. Despite this, there are few systematic comparisons of the structural behavior of these enzymes in various complexes. Here, (1)H-(15)N HSQC NMR spectroscopy is applied to PvuII endonuclease (2 x 18 kDa) in an effort to better understand the relationship between sequence recognition and enzyme conformational behavior. Spectra of the free enzyme collected in the absence and presence of metal ions indicate that while there is a modest backbone conformational response upon binding Ca(II), this does not occur with Mg(II). Substrate binding itself is accompanied by very dramatic spectral changes consistent with a large-scale conformational response. HSQC spectra of the enzyme bound to cognate (specific) and noncognate (nonspecific) oligonucleotides in the presence of Ca(II) are dramatically distinct, revealing for the first time the structural uniqueness of a PvuII cognate complex in solution. The strong correlation between NMR spectral overlap and crystallographic data (C(alpha) rmsd) permits characterization of the nonspecific PvuII complex as being more similar to the free enzyme than to the specific complex. Collectively, these data support the notion that it is the DNA, not the metal ion, which promotes a unique conformational response by the enzyme. It therefore follows that the principle role of metal ions in complex formation is one of driving substrate affinity and stability rather than conformationally priming the enzyme for substrate binding and sequence recognition. These results not only provide valuable insights into the mechanism of protein-DNA interactions but also demonstrate the utility of NMR spectroscopy in structure-function studies of these representative nucleic acid systems.

Regulation of transcription of genes

The principles of the mechanism of gene regulation as elucidated for prokaryotes apply in outline to the eukaryotic as well. For example, the mechanisms of protein-DNA interactions are similar and in some cases highly conserved throughout evolution. It is not difficult to imagine how control elements found in prokaryotes could have been combined during evolution to render any given gene in a eukaryote subject to multiple controls. Moreover, it is now clear that the best understood case of development — that of the early stages of Drosophila - may be described as a reiteration and integration of a series of ‘genetic switches’ similar to that found in, for example, the coliphage λ.

The significance of distance and orientation of restriction endonuclease recognition sites in viral DNA genomes

Studies on phage T3 and T7 have shown that these viruses avoid restriction not only by the phage-coded Ocr (and S-adenosylmethionine hydrolase) protein functions or by the complete loss of specific recognition sites for certain restriction endonucleases from their genomes, but also that there are two additional modes: resistance towards EcoP15 (which recognizes a non-symmetrical sequence) is achieved by an identical orientation of all the recognition sites in the virus genome (strand bias) and in the case of EcoRII by the extreme reduction in number and thereby greater distance between recognition sites in the genome. These observations led to the discovery that certain restriction endonucleases require the simultaneous cooperation with two DNA sites for their function, as well as to the ongoing elucidation of the molecular modes of action of these enzymes. Type II and type III enzymes display fundamentally different mechanisms of protein-DNA interaction. For EcoRII we favor a model of simultaneous binding of two DNA sites to a dimeric enzyme molecule (neighbouring sites of the same, looping, DNA molecule or sites located on different DNA molecules), while the action of EcoP15 seems to conform with a tracking-collision model of two enzyme molecules bound to inversely oriented recognition sites. In addition to podoviruses T3 and T7, strand bias of recognition sequences for different type III DNA modification-restriction enzymes is also observed in the inoviruses M13, IKE and PF3.

Surface plasmon resonance based kinetic studies of zinc finger-DNA interactions

Libraries of the zinc finger DNA binding protein, Zif268, have been constructed and selected for affinity and specificity toward DNA targets using the phage display technique (Wu et al., 1995). Mutant proteins were purified to homogeneity and were characterized for their ability to interact with their DNA targets using a real-time biomolecular interaction assay (BIA). One mutant protein, C7, bound the Zif268 consensus binding sequence with a 13-fold increase in affinity as compared to the wild-type Zif268 protein. Mutant proteins with moderate affinity for new DNA targets within a consensus sequence of HIV-1 have also been obtained. Surface plasmon resonance based BIA has provided invaluable kinetic information which offers insights into the mechanism of protein-DNA interactions.