Protein–DNA binding
Dotaz
Zobrazit nápovědu
1st ed. 427 s. : il.
- Klíčová slova
- DNA,
- MeSH
- DNA vazebné proteiny MeSH
- DNA MeSH
- genetické techniky MeSH
- Publikační typ
- příručky MeSH
Annals of the New York Academy of Sciences ; vol. 684
1st ed. IX, 258 s. : obr.,grafy, tab., přeruš.lit., věc.rejstř. ; 23 cm
- MeSH
- DNA vazebné proteiny MeSH
- molekulární biologie MeSH
- zinkové prsty MeSH
- Publikační typ
- kongresy MeSH
- Konspekt
- Patologie. Klinická medicína
- NLK Obory
- embryologie a teratologie
- biologie
- genetika, lékařská genetika
p53 plays critical roles in regulating cell cycle, apoptosis, senescence and metabolism and is commonly mutated in human cancer. These roles are achieved by interaction with other proteins, but particularly by interaction with DNA. As a transcription factor, p53 is well known to bind consensus target sequences in linear B-DNA. Recent findings indicate that p53 binds with higher affinity to target sequences that form cruciform DNA structure. Moreover, p53 binds very tightly to non-B DNA structures and local DNA structures are increasingly recognized to influence the activity of wild-type and mutant p53. Apart from cruciform structures, p53 binds to quadruplex DNA, triplex DNA, DNA loops, bulged DNA and hemicatenane DNA. In this review, we describe local DNA structures and summarize information about interactions of p53 with these structural DNA motifs. These recent data provide important insights into the complexity of the p53 pathway and the functional consequences of wild-type and mutant p53 activation in normal and tumor cells.
- MeSH
- B-DNA MeSH
- DNA chemie genetika metabolismus MeSH
- konformace nukleové kyseliny * MeSH
- lidé MeSH
- nádorový supresorový protein p53 chemie metabolismus MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
The histone-like DNA-binding proteins (HU) serve as model molecules for protein thermostability studies, as they function in different bacteria that grow in a wide range of temperatures and show sequence diversity under a common fold. In this work, we report the cloning of the hutth gene from Thermus thermophilus, the purification and crystallization of the recombinant HUTth protein, as well as its X-ray structure determination at 1.7 Å. Detailed structural and thermodynamic analyses were performed towards the understanding of the thermostability mechanism. The interaction of HUTth protein with plasmid DNA in solution has been determined for the first time with MST. Sequence conservation of an exclusively thermophilic order like Thermales, when compared to a predominantly mesophilic order (Deinococcales), should be subject, to some extent, to thermostability-related evolutionary pressure. This hypothesis was used to guide our bioinformatics and evolutionary studies. We discuss the impact of thermostability adaptation on the structure of HU proteins, based on the detailed evolutionary analysis of the Deinococcus-Thermus phylum, where HUTth belongs. Furthermore, we propose a novel method of engineering thermostable proteins, by combining consensus-based design with ancestral sequence reconstruction. Finally, through the structure of HUTth, we are able to examine the validity of these predictions. Our approach represents a significant advancement, as it explores for the first time the potential of ancestral sequence reconstruction in the divergence between a thermophilic and a mainly mesophilic taxon, combined with consensus-based engineering.
- MeSH
- bakteriální proteiny chemie genetika metabolismus MeSH
- DNA vazebné proteiny chemie genetika metabolismus MeSH
- konzervovaná sekvence MeSH
- molekulární evoluce * MeSH
- stabilita proteinů MeSH
- Thermus thermophilus genetika metabolismus MeSH
- vazba proteinů MeSH
- vysoká teplota * MeSH
- Publikační typ
- časopisecké články MeSH
Characterization of protein-protein and protein-DNA interactions is critical to understand mechanisms governing the biology of cells. Here we describe optimized methods and their mutual combinations for this purpose: bimolecular fluorescence complementation (BiFC), co-immunoprecipitation (Co-IP), yeast two-hybrid systems (Y2H), and chromatin immunoprecipitation (ChIP). These improved protocols detect trimeric complexes in which two proteins of interest interact indirectly via a protein sandwiched between them. They also allow isolation of low-abundance chromatin proteins and confirmation that proteins of interest are associated with specific DNA sequences, for example telomeric tracts. Here we describe these methods and their application to map interactions of several telomere- and telomerase-associated proteins and to purify a sufficient amount of chromatin from Arabidopsis thaliana for further investigations (e.g., next-generation sequencing, hybridization).
- MeSH
- Arabidopsis genetika metabolismus MeSH
- buněčné jádro metabolismus MeSH
- chromatin metabolismus MeSH
- chromatinová imunoprecipitace metody MeSH
- DNA rostlinná metabolismus MeSH
- DNA vazebné proteiny izolace a purifikace metabolismus MeSH
- imunoprecipitace metody MeSH
- mapování interakce mezi proteiny metody MeSH
- optické zobrazování metody MeSH
- proteiny huseníčku genetika metabolismus MeSH
- techniky dvojhybridového systému MeSH
- telomerasa metabolismus MeSH
- telomery metabolismus MeSH
- vazba proteinů MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The HMG-box domain of approximately 75 amino acid residues was originally identified as the domain that mediates the DNA-binding of chromatin-associated high-mobility group (HMG) proteins of the HMGB type. In the last few years, HMG-box domains have been found in various DNA-binding proteins including transcription factors and subunits of chromatin-remodeling complexes. HMG-box domains mediate either non-sequence-specific (e.g., HMGB-type proteins) or sequence-specific (e.g., transcription factors) DNA binding. Both types of HMG-box domains bind non-B-type DNA structures (bent, kinked and unwound) with high affinity. In addition, HMG-box domains are involved in a variety of protein-protein interactions. Here, we have examined the human and plant genomes for genes encoding HMG-box domains. Compared to plants, human cells contain a larger variety of HMG-box proteins. Whereas in humans transcription factors are the most divergent group of HMG-box proteins, in plants the chromosomal HMGB-type proteins are most variable.
- MeSH
- DNA vazebné proteiny MeSH
- domény HMG-Box MeSH
- financování organizované MeSH
- genom lidský MeSH
- genom rostlinný MeSH
- jaderné proteiny MeSH
- lidé MeSH
- proteiny Drosophily MeSH
- proteiny HMGB MeSH
- restrukturace chromatinu MeSH
- transkripční faktory MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- přehledy MeSH
The 14-3-3 protein family is a highly conserved and widely distributed group of proteins consisting of multiple isoforms in eukaryotes. Ubiquitously expressed, 14-3-3 proteins play key roles in DNA replication, cell cycle regulation, and apoptosis. The function of 14-3-3 proteins is mediated by interaction with a large number of other proteins and with DNA. It has been demonstrated that 14-3-3γ protein binds strongly to cruciform structures and is crucial for initiating replication. In this study, we analyzed DNA binding properties of the 14-3-3γ isoform to linear and supercoiled DNA. We demonstrate that 14-3-3γ protein binds strongly to long DNA targets, as evidenced by electrophoretic mobility shift assay on agarose gels. Binding of 14-3-3γ to DNA target results in the appearance of blurry, retarded DNA bands. Competition experiments with linear and supercoiled DNA on magnetic beads show very strong preference for supercoiled DNA. We also show by confocal microscopy that 14-3-3 protein in the HCT-116 cell line is co-localized with DNA cruciforms. This implies a role for the 14-3-3γ protein in its binding to local DNA structures which are stabilized by DNA supercoiling.
- MeSH
- Escherichia coli genetika MeSH
- HCT116 buňky MeSH
- klonování DNA MeSH
- kompetitivní vazba MeSH
- křížová struktura DNA genetika metabolismus MeSH
- lidé MeSH
- plazmidy genetika MeSH
- proteiny 14-3-3 genetika metabolismus MeSH
- rekombinantní fúzní proteiny genetika metabolismus MeSH
- replikace DNA genetika MeSH
- retardační test MeSH
- superhelikální DNA genetika metabolismus MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH