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1.
J Chem Phys ; 151(12): 125101, 2019 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-31575173

RESUMO

Gene regulation is one of the most important fundamental biological processes in living cells. It involves multiple protein molecules that locate specific sites on DNA and assemble gene initiation or gene repression multimolecular complexes. While the protein search dynamics for DNA targets has been intensively investigated, the role of intermolecular interactions during the genetic activation or repression remains not well quantified. Here, we present a simple one-dimensional model of target search for two interacting molecules that can reversibly form a dimer molecular complex, which also participates in the search process. In addition, the proteins have finite residence times on specific target sites, and the gene is activated or repressed when both proteins are simultaneously present at the target. The model is analyzed using first-passage analytical calculations and Monte Carlo computer simulations. It is shown that the search dynamics exhibit a complex behavior depending on the strength of intermolecular interactions and on the target residence times. We also found that the search time shows a nonmonotonic behavior as a function of the dissociation rate for the molecular complex. Physical-chemical arguments to explain these observations are presented. Our theoretical approach highlights the importance of molecular interactions in the complex process of gene activation/repression by multiple transcription factor proteins.


Assuntos
DNA/química , Modelos Químicos , Simulação por Computador , DNA/genética , DNA/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Cinética , Método de Monte Carlo , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
3.
Inorg Chem ; 58(17): 11351-11363, 2019 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-31433627

RESUMO

Iron storage in biology is carried out by cage-shaped proteins of the ferritin superfamily, one of which is the dodecameric protein Dps. In Dps, four distinct steps lead to the formation of metal nanoparticles: attraction of ion-aquo complexes to the protein matrix, passage of these complexes through translocation pores, oxidation of these complexes at ferroxidase centers, and, ultimately, nanoparticle formation. In this study, we investigated Dps from Listeria innocua to structurally characterize these steps for Co2+, Zn2+, and La3+ ions. The structures reveal that differences in their ion coordination chemistry determine alternative metal ion-binding sites on the areas of the surface surrounding the translocation pore that captures nine La3+, three Co2+, or three Zn2+ ions as aquo clusters and passes them on for translocation. Inside these pores, ion-selective conformational changes at key residues occur before a gating residue to actively move ions through the constriction zone. Ions upstream of the Asp130 gate residue are typically hydrated, while ions downstream directly interact with the protein matrix. Inside the cavity, ions move along negatively charged residues to the ferroxidase center, where seven main residues adapt to the three different ions by dynamically changing their conformations. In total, we observed more than 20 metal-binding sites per Dps monomer, which clearly highlights the metal-binding capacity of this protein family. Collectively, our results provide a detailed structural description of the preparative steps for amino acid-assisted biomineralization in Dps proteins, demonstrating unexpected protein matrix plasticity.


Assuntos
Proteínas de Bactérias/química , Proteínas de Ligação a DNA/química , Listeria/química , Metais Pesados/química , Proteínas de Bactérias/biossíntese , Proteínas de Ligação a DNA/biossíntese , Modelos Moleculares , Eletricidade Estática
4.
Yakugaku Zasshi ; 139(7): 969-973, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31257254

RESUMO

Translesion DNA synthesis (TLS) is an emergency system activated to inhibit cell death caused by DNA damage-induced replication arrest. Thus, TLS enables cancer cells to acquire resistance to alkylate anticancer drugs. REV7 functions as the hub protein that interacts with both the inserter DNA polymerase REV1 and the extender DNA polymerase REV3 in TLS. REV7-mediated protein-protein interactions (PPIs) are essential for the activation of TLS, and are therefore attractive targets for anticancer drug development. To clarify the REV7-REV3 and REV7-REV1 PPIs, we determined the structures of REV7-REV3 and REV7-REV3-REV1 complexes. In the structures of REV7-REV3 and REV7-REV3-REV1 complexes, REV7 wraps around the REV3 fragment, and the REV1-binding interface is distinct from the REV3-binding site of REV7. We also identified a novel REV7 binding protein, transcription factor II-I (TFII-I), which is required for TLS. Of note, TFII-I binds the REV7-REV3-REV1 complex, suggesting that REV7-TFII-I PPIs are independent of other REV7-mediated PPIs. Furthermore, we found a small-molecule compound that inhibits TLS by targeting the REV7-REV3 PPIs. Lastly, we determined the structure of REV7 in complex with chromosome alignment maintaining phosphoprotein (CAMP), a known kinetochore-microtubule attachment protein. The overall structure of the REV7-CAMP complex is similar to that of the REV7-REV3 complex, but the REV7-CAMP PPIs are markedly different from the REV7-REV3 PPIs. These findings improve our understanding of multifunctional hub proteins, and are helpful for designing small-molecule compounds for novel anticancer drug development.


Assuntos
Antineoplásicos , Descoberta de Drogas , Proteína Semelhante a ELAV 2/química , Cristalografia por Raios X , DNA/biossíntese , Dano ao DNA , Proteínas de Ligação a DNA/química , DNA Polimerase Dirigida por DNA/química , Humanos , Proteínas Mad2/química , Peso Molecular , Proteínas Nucleares/química , Nucleotidiltransferases/química , Ligação Proteica , Mapas de Interação de Proteínas , Estrutura Terciária de Proteína
5.
J Chem Theory Comput ; 15(8): 4687-4698, 2019 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-31251056

RESUMO

Small-angle X-ray and small-angle neutron scattering (SAXS/SANS) provide unique structural information on biomolecules and their complexes in solution. SANS may provide multiple independent data sets by means of contrast variation experiments, that is, by measuring at different D2O concentrations and different perdeuteration conditions of the biomolecular complex. However, even the combined data from multiple SAXS/SANS sets is by far insufficient to define all degrees of freedom of a complex, leading to a significant risk of overfitting when refining biomolecular structures against SAXS/SANS data. Hence, to control against overfitting, the low-information SAXS/SANS data must be complemented by accurate physical models, and, if possible, refined models should be cross-validated against independent data not used during the refinement. We present a method for refining atomic biomolecular structures against multiple sets of SAXS and SANS data using all-atom molecular dynamics simulations. Using the protein citrate synthase and the protein/RNA complex Sxl-Unr-msl2 mRNA as test cases, we demonstrate how multiple SAXS and SANS sets may be used for refinement and cross-validation, thereby excluding overfitting during refinement. For the Sxl-Unr-msl2 complex, we find that perdeuteration of the Unr domain leads to a unique, slightly compacted conformation, whereas other perdeuteration conditions lead to similar solution conformations compared to the nondeuterated state. In line with our previous method for predicting SAXS curves, SANS curves were predicted with explicit-solvent calculations, taking atomic models for both the hydration layer and the excluded solvent into account, thereby avoiding the use of solvent-related fitting parameters and solvent-reduced neutron scattering lengths. We expect the method to be useful for deriving and validating solution structures of biomolecules and soft-matter complexes, and for critically assessing whether multiple SAXS and SANS sets are mutually compatible.


Assuntos
Simulação de Dinâmica Molecular , Difração de Nêutrons , Espalhamento a Baixo Ângulo , Difração de Raios X , Animais , Citrato (si)-Sintase/química , Proteínas de Ligação a DNA/química , Drosophila , Proteínas de Drosophila/química , Difração de Nêutrons/métodos , RNA/química , Proteínas de Ligação a RNA/química , Suínos , Difração de Raios X/métodos
6.
Chem Commun (Camb) ; 55(52): 7466-7469, 2019 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-31184647
7.
Subcell Biochem ; 92: 169-186, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31214987

RESUMO

The periplasm of Gram-negative bacteria contains a specialized chaperone network that facilitates the transport of unfolded membrane proteins to the outer membrane as its primary functional role. The network, involving the chaperones Skp and SurA as key players and potentially additional chaperones, is indispensable for the survival of the cell. Structural descriptions of the apo forms of these molecular chaperones were initially provided by X-ray crystallography. Subsequently, a combination of experimental biophysical methods including solution NMR spectroscopy provided a detailed understanding of full-length chaperone-client complexes . The data showed that conformational changes and dynamic re-organization of the chaperones upon client binding, as well as client dynamics on the chaperone surface are crucial for function. This chapter gives an overview of the structure-function relationship of the dynamic conformational rearrangements that regulate the functional cycles of the periplasmic molecular chaperones Skp and SurA.


Assuntos
Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Bactérias Gram-Negativas/metabolismo , Chaperonas Moleculares/química , Chaperonas Moleculares/metabolismo , Peptidilprolil Isomerase/química , Peptidilprolil Isomerase/metabolismo , Periplasma/metabolismo , Bactérias Gram-Negativas/enzimologia
8.
Nat Commun ; 10(1): 2104, 2019 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-31068591

RESUMO

Protein-induced fluorescence enhancement (PIFE) is a popular tool for characterizing protein-DNA interactions. PIFE has been explained by an increase in local viscosity due to the presence of the protein residues. This explanation, however, denies the opposite effect of fluorescence quenching. This work offers a perspective for understanding PIFE mechanism and reports the observation of a phenomenon that we name protein-induced fluorescence quenching (PIFQ), which exhibits an opposite effect to PIFE. A detailed characterization of these two fluorescence modulations reveals that the initial fluorescence state of the labeled mediator (DNA) determines whether this mediator-conjugated dye undergoes PIFE or PIFQ upon protein binding. This key role of the mediator DNA provides a protocol for the experimental design to obtain either PIFQ or PIFE, on-demand. This makes the arbitrary nature of the current experimental design obsolete, allowing for proper integration of both PIFE and PIFQ with existing bulk and single-molecule fluorescence techniques.


Assuntos
DNA/metabolismo , Corantes Fluorescentes/química , Imagem Individual de Molécula/métodos , DNA/química , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/isolamento & purificação , Proteínas de Ligação a DNA/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/isolamento & purificação , Proteínas de Escherichia coli/metabolismo , Endonucleases Flap/química , Endonucleases Flap/isolamento & purificação , Endonucleases Flap/metabolismo , Fluorescência , Transferência Ressonante de Energia de Fluorescência/métodos , Microscopia de Fluorescência/métodos , Oligonucleotídeos/química , Oligonucleotídeos/metabolismo , Ligação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Coloração e Rotulagem , Proteínas Virais/química , Proteínas Virais/isolamento & purificação , Proteínas Virais/metabolismo
9.
Nat Commun ; 10(1): 2314, 2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-31127101

RESUMO

Histone methyltransferase MLL4 is centrally involved in transcriptional regulation and is often mutated in human diseases, including cancer and developmental disorders. MLL4 contains a catalytic SET domain that mono-methylates histone H3K4 and seven PHD fingers of unclear function. Here, we identify the PHD6 finger of MLL4 (MLL4-PHD6) as a selective reader of the epigenetic modification H4K16ac. The solution NMR structure of MLL4-PHD6 in complex with a H4K16ac peptide along with binding and mutational analyses reveal unique mechanistic features underlying recognition of H4K16ac. Genomic studies show that one third of MLL4 chromatin binding sites overlap with H4K16ac-enriched regions in vivo and that MLL4 occupancy in a set of genomic targets depends on the acetyltransferase activity of MOF, a H4K16ac-specific acetyltransferase. The recognition of H4K16ac is conserved in the PHD7 finger of paralogous MLL3. Together, our findings reveal a previously uncharacterized acetyllysine reader and suggest that selective targeting of H4K16ac by MLL4 provides a direct functional link between MLL4, MOF and H4K16 acetylation.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Histona Acetiltransferases/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Dedos de Zinco PHD/fisiologia , Acetilação , Animais , Sítios de Ligação , Cromatina/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/isolamento & purificação , Técnicas de Inativação de Genes , Células HEK293 , Histona Acetiltransferases/genética , Histona-Lisina N-Metiltransferase/química , Histonas/química , Humanos , Camundongos Transgênicos , Modelos Moleculares , Mutagênese Sítio-Dirigida , Ressonância Magnética Nuclear Biomolecular , Processamento de Proteína Pós-Traducional/fisiologia , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo
10.
Anal Bioanal Chem ; 411(16): 3581-3589, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31089784

RESUMO

The use of aptamers in various analytical applications as molecular recognition elements and alternative to antibodies has led to the development of various platforms that facilitate the sensitive and specific detection of targets ranging from small molecules and proteins to whole cells. The goal of this work was to design a universal and adaptable sandwich-type aptasensor exploiting the unique properties of DNA binding proteins. Specifically, two different enzyme-DNA binding protein conjugates, GOx-dHP and HRP-scCro, were used for the direct detection of a protein using two aptamers for target capture and detection. The specific dsDNA binding sequence for each DNA binding protein tag was incorporated in the form of a hairpin at one end of each aptamer sequence during the synthesis step. Detection was accomplished by an enzymatic (GOx/HRP) cascade reaction after the binding of each enzyme conjugate to its corresponding binding sequence on each aptamer. The proposed sandwich-type aptasensor was validated for the detection of thrombin, which is one of the most commonly used model targets with known dual aptamers. The limit of detection accomplished was 0.92 nM which is comparable with other colorimetric platforms reported in the literature. The sensitivity of the aptasensor was easily modulated by changing the number of dsDNA binding sites incorporated in the aptamer sequences, thus controlling the enzyme stoichiometry. Finally, the potential use of the proposed sensing approach for real sample testing was demonstrated using spiked human plasma and no significant matrix effects were observed when up to 2% plasma was used.


Assuntos
Aptâmeros de Nucleotídeos/química , Proteínas de Ligação a DNA/química , Glucose Oxidase/química , Peroxidase do Rábano Silvestre/química , Sequência de Bases , Sítios de Ligação , Técnicas Biossensoriais , Proteínas Sanguíneas/análise , Colorimetria/métodos , Técnicas Eletroquímicas/métodos , Ensaio de Desvio de Mobilidade Eletroforética , Humanos , Limite de Detecção , Reprodutibilidade dos Testes
11.
Molecules ; 24(9)2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-31067825

RESUMO

G-quadruplex (G4) structures are highly stable four-stranded DNA and RNA secondary structures held together by non-canonical guanine base pairs. G4 sequence motifs are enriched at specific sites in eukaryotic genomes, suggesting regulatory functions of G4 structures during different biological processes. Considering the high thermodynamic stability of G4 structures, various proteins are necessary for G4 structure formation and unwinding. In a yeast one-hybrid screen, we identified Slx9 as a novel G4-binding protein. We confirmed that Slx9 binds to G4 DNA structures in vitro. Despite these findings, Slx9 binds only insignificantly to G-rich/G4 regions in Saccharomyces cerevisiae as demonstrated by genome-wide ChIP-seq analysis. However, Slx9 binding to G4s is significantly increased in the absence of Sgs1, a RecQ helicase that regulates G4 structures. Different genetic and molecular analyses allowed us to propose a model in which Slx9 recognizes and protects stabilized G4 structures in vivo.


Assuntos
Proteínas de Ligação a DNA/química , Quadruplex G , Proteínas Ribossômicas/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , DNA Helicases/química , DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Genoma/genética , Conformação de Ácido Nucleico , Ligação Proteica , RecQ Helicases/química , RecQ Helicases/genética , Proteínas Ribossômicas/química , Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/química , Termodinâmica
12.
Nat Commun ; 10(1): 1960, 2019 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-31036859

RESUMO

Lin28-dependent oligo-uridylylation of precursor let-7 (pre-let-7) by terminal uridylyltransferase 4/7 (TUT4/7) represses let-7 expression by blocking Dicer processing, and regulates cell differentiation and proliferation. The interaction between the Lin28:pre-let-7 complex and the N-terminal Lin28-interacting module (LIM) of TUT4/7 is required for pre-let-7 oligo-uridylylation by the C-terminal catalytic module (CM) of TUT4/7. Here, we report crystallographic and biochemical analyses of the LIM of human TUT4. The LIM consists of the N-terminal Cys2His2-type zinc finger (ZF) and the non-catalytic nucleotidyltransferase domain (nc-NTD). The ZF of LIM adopts a distinct structural domain, and its structure is homologous to those of double-stranded RNA binding zinc fingers. The interaction between the ZF and pre-let-7 stabilizes the Lin28:pre-let-7:TUT4 ternary complex, and enhances the oligo-uridylylation reaction by the CM. Thus, the ZF in LIM and the zinc-knuckle in the CM, which interacts with the oligo-uridylylated tail, together facilitate Lin28-dependent pre-let-7 oligo-uridylylation.


Assuntos
Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , MicroRNAs/metabolismo , RNA Nucleotidiltransferases/química , RNA Nucleotidiltransferases/metabolismo , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Cristalografia por Raios X , Proteínas de Ligação a DNA/genética , Humanos , MicroRNAs/genética , Ligação Proteica , RNA Nucleotidiltransferases/genética , Proteínas de Ligação a RNA/genética
14.
Chem Biol Interact ; 308: 244-251, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31145890

RESUMO

Increasing evidence has shown that dysregulation of microRNA-621 (miR-621) is demonstrated to be associated with several cancers. However, the role of miR-621 in bladder cancer (BCa) remains unclear. Herein, we aimed to study the expression pattern, biological function, and molecular mechanism of miR-621 in BCa. First, we demonstrated that miR-621 was frequently downregulated in BCa tissues and cell lines compared with the adjacent normal BCa tissues and non-cancerous immortalized urothelial cell line. In addition, the expression of miR-621 was negatively correlated with overall survival of BCa patients. Functional experiments suggessted that miR-621 inhibited the proliferation and metastasis of BCa cells. Notably, dual-luciferase assay showed that miR-621 directly targeted the 3' UTR of TRIM29, which was frequently upregulated in BCa tissues and displayed inverse correlation with miR-621 expression. Furthermore, we demonstrated that miR-621 inhibited the proliferation and metastasis of BCa cells via Wnt/ß-catenin signaling pathway by targeting TRIM29. Our study suggested that the miR-621/TRIM29 axis inhibits the proliferation and metastasis of BCa cells via Wnt/ß-catenin signaling pathway and may have potential applications for development of BCa diagnosis or treatment.


Assuntos
Proliferação de Células , MicroRNAs/metabolismo , Via de Sinalização Wnt , Regiões 3' não Traduzidas , Antagomirs/metabolismo , Sequência de Bases , Linhagem Celular Tumoral , Movimento Celular , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Alinhamento de Sequência , Fatores de Transcrição/química , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologia
15.
Gene ; 710: 48-58, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31145962

RESUMO

We evaluated microsatellite instability (MSI) in selected mismatch repair (MMR) and tumor suppressor (TS) genes with a view to exploring genetic changes associated with the occurrence of gastric cancer (GC). Moreover, expression of MSI positive genes was measured to get insights into molecular events operating in the tumor microenvironment. We anticipated discovering new molecular targets with potential as molecular biomarkers of gastric cancer. Of the 13 genes screened, we observed 15% to 52.5% MSI at eight microsatellite loci located in 3' UTR and coding regions of six genes (TGFBR2, PDCD4, MLH3, DLC1, MSH6, and MSH3). The union probability of different combinations of unstable microsatellite loci unveiled a set of four MSI markers from TGFBR2, PDCD4, MLH3, and MSH3 genes that allows detection of up to 85% incidences of GC. Significant downregulation of MLH3, PDCD4, TGFBR2, and DLC1 genes was observed in tumor tissues. Protein structure analyses of two unexplored targets, MSH3 (TG4) and MSH6 (A7), with MSI in the coding region, exhibited the loss of essential domains in the encoded aberrant protein hampering its function in the MMR machinery. The molecular markers thus identified could potentially be used as MSI biomarkers for the diagnosis of gastric tumorigenesis after further validation.


Assuntos
Biomarcadores Tumorais/genética , Perfilação da Expressão Gênica/métodos , Instabilidade de Microssatélites , Neoplasias Gástricas/genética , Regiões 3' não Traduzidas , Reparo de Erro de Pareamento de DNA , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Detecção Precoce de Câncer , Regulação Neoplásica da Expressão Gênica , Humanos , Proteína 3 Homóloga a MutS/química , Proteína 3 Homóloga a MutS/genética , Estrutura Terciária de Proteína , Microambiente Tumoral , Proteínas Supressoras de Tumor/genética
16.
Nature ; 571(7763): 79-84, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31142837

RESUMO

Access to DNA packaged in nucleosomes is critical for gene regulation, DNA replication and DNA repair. In humans, the UV-damaged DNA-binding protein (UV-DDB) complex detects UV-light-induced pyrimidine dimers throughout the genome; however, it remains unknown how these lesions are recognized in chromatin, in which nucleosomes restrict access to DNA. Here we report cryo-electron microscopy structures of UV-DDB bound to nucleosomes bearing a 6-4 pyrimidine-pyrimidone dimer or a DNA-damage mimic in various positions. We find that UV-DDB binds UV-damaged nucleosomes at lesions located in the solvent-facing minor groove without affecting the overall nucleosome architecture. In the case of buried lesions that face the histone core, UV-DDB changes the predominant translational register of the nucleosome and selectively binds the lesion in an accessible, exposed position. Our findings explain how UV-DDB detects occluded lesions in strongly positioned nucleosomes, and identify slide-assisted site exposure as a mechanism by which high-affinity DNA-binding proteins can access otherwise occluded sites in nucleosomal DNA.


Assuntos
Dano ao DNA , Proteínas de Ligação a DNA/metabolismo , DNA/metabolismo , DNA/ultraestrutura , Nucleossomos/metabolismo , Nucleossomos/ultraestrutura , Dímeros de Pirimidina/análise , Microscopia Crioeletrônica , DNA/química , DNA/efeitos da radiação , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/ultraestrutura , Histonas/química , Histonas/metabolismo , Histonas/ultraestrutura , Humanos , Modelos Moleculares , Nucleossomos/genética , Nucleossomos/efeitos da radiação , Dímeros de Pirimidina/química , Dímeros de Pirimidina/genética , Termodinâmica , Raios Ultravioleta/efeitos adversos
17.
BMC Med Genet ; 20(1): 79, 2019 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-31077156

RESUMO

BACKGROUND: The X-ray repair cross-complementing group 3 (XRCC3) is an efficient component of homologous recombination and is required for the preservation of chromosomal integrity in mammalian cells. The association between Thr241Met single-nucleotide polymorphism (SNP) in this gene and susceptibility to breast cancer has been assessed in several studies. Yet, reports are controversial. The present meta-analysis has been designed to identify whether this SNP is associated with susceptibility to breast cancer. METHODS: We performed a systematic review and meta-analysis for retrieving the case-control studies on the associations between T241 M SNP and the risk of breast cancer. Crude odds ratios (ORs) and 95% confidence intervals (CIs) were calculated to verify the association in dominant, recessive, and homozygote inheritance models. RESULTS: We included 55 studies containing 30,966 sporadic breast cancer cases, 1174 familial breast cancer cases and 32,890 controls in the meta-analysis. In crude analyses, no association was detected between the mentioned SNP and breast cancer risk in recessive, homozygote or dominant models. However, ethnic based analysis showed that in sporadic breast cancer, the SNP was associated with breast cancer risk in Arab populations in homozygous (OR (95% CI) = 3.649 (2.029-6.563), p = 0.0001) and recessive models (OR (95% CI) = 4.092 (1.806-9.271), p = 0.001). The association was significant in Asian population in dominant model (OR (95% CI) = 1.296, p = 0.029). However, the associations was significant in familial breast cancer in mixed ethnic-based subgroup in homozygote and recessive models (OR (95% CI) = 0.451 (0.309-0.659), p = 0.0001, OR (95% CI) = 0.462 (0.298-0.716), p = 0.001 respectively). CONCLUSIONS: Taken together, our results in a large sample of both sporadic and familial cases of breast cancer showed insignificant role of Thr241Met in the pathogenesis of this type of malignancy. Such results were more conclusive in sporadic cases. In familial cases, future studies are needed to verify our results.


Assuntos
Neoplasias da Mama/genética , Proteínas de Ligação a DNA/genética , Predisposição Genética para Doença , Metionina/genética , Treonina/genética , Proteínas de Ligação a DNA/química , Feminino , Recombinação Homóloga , Humanos , Polimorfismo de Nucleotídeo Único
18.
PLoS Comput Biol ; 15(4): e1006768, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30933978

RESUMO

Recognition of single-stranded DNA (ssDNA) or single-stranded RNA (ssRNA) is important for many fundamental cellular functions. A variety of single-stranded DNA-binding proteins (ssDBPs) and single-stranded RNA-binding proteins (ssRBPs) have evolved that bind ssDNA and ssRNA, respectively, with varying degree of affinities and specificities to form complexes. Structural studies of these complexes provide key insights into their recognition mechanism. However, computational modeling of the specific recognition process and to predict the structure of the complex is challenging, primarily due to the heterogeneity of their binding energy landscape and the greater flexibility of ssDNA or ssRNA compared with double-stranded nucleic acids. Consequently, considerably fewer computational studies have explored interactions between proteins and single-stranded nucleic acids compared with protein interactions with double-stranded nucleic acids. Here, we report a newly developed energy-based coarse-grained model to predict the structure of ssDNA-ssDBP and ssRNA-ssRBP complexes and to assess their sequence-specific interactions and stabilities. We tuned two factors that can modulate specific recognition: base-aromatic stacking strength and the flexibility of the single-stranded nucleic acid. The model was successfully applied to predict the binding conformations of 12 distinct ssDBP and ssRBP structures with their cognate ssDNA and ssRNA partners having various sequences. Estimated binding energies agreed well with the corresponding experimental binding affinities. Bound conformations from the simulation showed a funnel-shaped binding energy distribution where the native-like conformations corresponded to the energy minima. The various ssDNA-protein and ssRNA-protein complexes differed in the balance of electrostatic and aromatic energies. The lower affinity of the ssRNA-ssRBP complexes compared with the ssDNA-ssDBP complexes stems from lower flexibility of ssRNA compared to ssDNA, which results in higher rate constants for the dissociation of the complex (koff) for complexes involving the former.


Assuntos
DNA de Cadeia Simples/química , DNA de Cadeia Simples/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , RNA/química , RNA/metabolismo , Animais , Sequência de Bases , Biologia Computacional , Humanos , Modelos Moleculares , Conformação de Ácido Nucleico , Ligação Proteica , Conformação Proteica , Dobramento de Proteína , Domínios e Motivos de Interação entre Proteínas , Eletricidade Estática , Termodinâmica
19.
mSphere ; 4(2)2019 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-30971446

RESUMO

Most urinary tract infections (UTIs) are caused by uropathogenic Escherichia coli (UPEC), which depends on an extracellular organelle (type 1 pili) for adherence to bladder cells during infection. Type 1 pilus expression is partially regulated by inversion of a piece of DNA referred to as fimS, which contains the promoter for the fim operon encoding type 1 pili. fimS inversion is regulated by up to five recombinases collectively known as Fim recombinases. These Fim recombinases are currently known to regulate two other switches: the ipuS and hyxS switches. A long-standing question has been whether the Fim recombinases regulate the inversion of other switches, perhaps to coordinate expression for adhesion or virulence. We answered this question using whole-genome sequencing with a newly developed algorithm (structural variation detection using relative entropy [SVRE]) for calling structural variations using paired-end short-read sequencing. SVRE identified all of the previously known switches, refining the specificity of which recombinases act at which switches. Strikingly, we found no new inversions that were mediated by the Fim recombinases. We conclude that the Fim recombinases are each highly specific for a small number of switches. We hypothesize that the unlinked Fim recombinases have been recruited to regulate fimS, and fimS only, as a secondary locus; this further implies that regulation of type 1 pilus expression (and its role in gastrointestinal and/or genitourinary colonization) is important enough, on its own, to influence the evolution and maintenance of multiple additional genes within the accessory genome of E. coli IMPORTANCE UTI is a common ailment that affects more than half of all women during their lifetime. The leading cause of UTIs is UPEC, which relies on type 1 pili to colonize and persist within the bladder during infection. The regulation of type 1 pili is remarkable for an epigenetic mechanism in which a section of DNA containing a promoter is inverted. The inversion mechanism relies on what are thought to be dedicated recombinase genes; however, the full repertoire for these recombinases is not known. We show here that there are no additional targets beyond those already identified for the recombinases in the entire genome of two UPEC strains, arguing that type 1 pilus expression itself is the driving evolutionary force for the presence of these recombinase genes. This further suggests that targeting the type 1 pilus is a rational alternative nonantibiotic strategy for the treatment of UTI.


Assuntos
Proteínas de Fímbrias/genética , Fímbrias Bacterianas/genética , Regulação Bacteriana da Expressão Gênica , Inversão de Sequência , Escherichia coli Uropatogênica/genética , Algoritmos , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Entropia , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Fímbrias/química , Humanos , Integrases/química , Integrases/genética , Regiões Promotoras Genéticas , Infecções Urinárias/microbiologia , Virulência/genética
20.
Essays Biochem ; 63(1): 147-156, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-30940742

RESUMO

Chromatin comprises proteins, DNA and RNA, and its function is to condense and package the genome in a way that allows the necessary transactions such as transcription, replication and repair to occur in a highly organised and regulated manner. The packaging of chromatin is often thought of in a hierarchical fashion starting from the most basic unit of DNA packaging, the nucleosome, to the condensation of nucleosomal 'beads on a string' by linker histones to form the 30-nm fibre and eventually large chromatin domains. However, a picture of a more heterogeneous, dynamic and liquid-like assembly is emerging, in which intrinsically disordered proteins (IDPs) and proteins containing intrinsically disordered regions (IDRs) play a central role. Disorder features at all levels of chromatin organisation, from the histone tails, which are sites of extensive post-translational modification (PTM) that change the fate of the underlying genomic information, right through to transcription hubs, and the recently elucidated roles of IDPs and IDRs in the condensation of large regions of the genome through liquid-liquid phase separation.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas Intrinsicamente Desordenadas/metabolismo , Nucleossomos/metabolismo , Animais , Proteínas de Ligação a DNA/química , Humanos , Proteínas Intrinsicamente Desordenadas/química , Nucleossomos/química , Ligação Proteica , Domínios Proteicos , Processamento de Proteína Pós-Traducional
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