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1.
Preferential affinity of high molecular weight high mobility group non-histone chromatin proteins for single-stranded DNA. 总被引:4,自引:0,他引:4
P J Isackson J L Fishback D L Bidney G R Reeck 《The Journal of biological chemistry》1979,254(13):5569-5572
We have subjected proteins dissociated from chicken erythrocyte or calf thymus chromatin by 0.35 M NaCl to sequential chromatography on columns containing immobilized double-stranded DNA and single-stranded DNA. At 0.2 M NaCl, 1 mM Tris . Cl (pH 7.5), the high molecular weight, high mobility group proteins (HMG-1, HMG-2, and HMG-E), were not retained by double-stranded DNA columns, but were retained by single-stranded DNA columns. Thus, in that solvent, those proteins exhibit selective affinity for single-stranded DNA. This suggests that the functions of the high molecular weight, high mobility group proteins might involve destabilizing the DNA double helix by virtue of their preferential affinity for single-stranded DNA. 相似文献
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Cytoplasmic DNA-binding proteins 总被引:1,自引:0,他引:1
Cytoplasmic DNA-binding proteins were isolated from Chinese hamster liver, kidney and tissue culture cells by DNA-polyacrylamide chromatography. With homologous Chinese hamster DNA, and with calf thymus DNA, 1.4% of the proteins were bound to the column. With single-stranded DNA and with heterologous Micrococcus lysodeikticus DNA there was only 0.3% binding, suggesting the proteins preferentially bind to double-stranded DNA and show some sequence specificity. By a nitrocellulose filter assay the bound proteins had at least a 4- to 7-fold greater affinity for DNA than bulk cytoplasmic protein. SDS gel electrophoresis showed that specific proteins were being markedly concentrated by the column and it was primarily the high molecular weight proteins of 65 000 D and over which showed sequence specificity. Some proteins appeared in common with different organs, others were unique. These studies thus define a group of high molecular weight, cytoplasmic proteins which bind to native DNA with a degree of sequence specificity. Their possible relationship to gene regulation is discussed. 相似文献
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Dana Vuzman 《Journal of molecular biology》2010,396(3):674-684
The search through nonspecific DNA for a specific site by proteins is known to be facilitated by sliding, hopping, and intersegment transfer between separate DNA strands, yet the driving forces of these protein dynamics from the molecular perspective are unclear. In this study, molecular features of the DNA search mechanism were explored for three homologous proteins (the HoxD9, Antp, and NK-2 homeodomains) using a simple computational model in which protein-DNA interactions are represented solely by electrostatic forces. In particular, we studied the impact that disordered N-terminal tails (N-tails), which are more common in DNA-binding proteins than in other proteins, have on the efficiency of DNA search. While the three homeodomain proteins were found to use similar binding interfaces in specific and nonspecific interactions with DNAs, their different electrostatic potentials affect the nature of their sliding dynamics. The different lengths and net charges of the N-tails of the homeodomains affect their motion along the DNA. The presence of an N-tail increases sliding propensity but slows linear diffusion along the DNA. When the search is performed in the presence of two parallel DNA molecules, a direct transfer, which is facilitated by the protein tail, from one nonspecific DNA to another occurs. The tailed proteins jump between two DNA molecules through an intermediate in which the recognition helix of the protein is adsorbed to one DNA fragment and the N-tail is adsorbed to the second, suggesting a “monkey bar” mechanism. Our study illustrates how the molecular architecture of proteins controls the efficiency of DNA scanning. 相似文献
5.
The past decade has witnessed an exciting evolution in our understanding of eukaryotic DNA replication at the molecular level. Progress has been particularly rapid within the last few years due to the convergence of research on a variety of cell types, from yeast to human, encompassing disciplines ranging from clinical immunology to the molecular biology of viruses. New eukaryotic DNA replicases and accessory proteins have been purified and characterized, and some have been cloned and sequenced. In vitro systems for the replication of viral DNA have been developed, allowing the identification and purification of several mammalian replication proteins. In this review we focus on DNA polymerases alpha and delta and the polymerase accessory proteins, their physical and functional properties, as well as their roles in eukaryotic DNA replication. 相似文献
6.
Multiple species of single-stranded nucleic acid-binding proteins in Saccharomyces cerevisiae 总被引:9,自引:0,他引:9
DNA affinity chromatography has been used to identify the major single-stranded nucleic acid binding proteins (SSBs) of Saccharomyces cerevisiae. There are five abundant species having molecular masses of 50, 45, 31, 23, and 20 kDa. Four of these proteins are cytoplasmic and one is mitochondrial. To date, three of the proteins have been purified to homogeneity. The purified proteins are designated SSB-m, SSB-1, and SSB-2, with molecular masses of 20, 45, and 50 kDa, respectively. SSB-m is found only in mitochondrial subcellular fractions. SSB-1 stimulates purified yeast DNA polymerase I, while SSB-2 inhibits DNA polymerase I. An antibody against SSB-1 has been prepared in rabbits and purified by SSB-1-Sepharose affinity chromatography. The purified antibody specifically inhibits DNA synthesis in an in vitro replication system, suggesting that SSB-1 may be involved in DNA replication in vivo. SSB-2 has the highest affinity for single-stranded DNA of all three proteins. It may represent a new class of eukaryotic SSB, on the basis of molecular weight, inhibition of DNA polymerase and antigenicity. Antibodies have also been prepared against SSB-2. The immunological reagents have been used to show that SSB-1, SSB-2, and SSB-m are antigenically distinct, as well as to study the relationship of these three SSBs to other proteins in yeast. 相似文献
7.
Conjugation is a paradigmatic example of horizontal or lateral gene transfer, whereby DNA is translocated between bacterial cells. It provides a route for the rapid acquisition of new genetic information. Increased antibiotic resistance among pathogens is a troubling consequence of this microbial capacity. DNA transfer across cell membranes requires a sophisticated molecular machinery that involves the participation of several proteins in DNA processing and replication, cell recruitment, and the transport of DNA and proteins from donor to recipient cells. Although bacterial conjugation was first reported in the 1940s, only now are we beginning to unravel the molecular mechanisms behind this process. In particular, structural biology is revealing the detailed molecular architecture of several of the pieces involved. 相似文献
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Recent work revealed a new class of molecular machines called molecular sleds, which are small basic molecules that bind and slide along DNA with the ability to carry cargo along DNA. Here, we performed biochemical and single-molecule flow stretching assays to investigate the basis of sliding activity in molecular sleds. In particular, we identified the functional core of pVIc, the first molecular sled characterized; peptide functional groups that control sliding activity; and propose a model for the sliding activity of molecular sleds. We also observed widespread DNA binding and sliding activity among basic polypeptide sequences that implicate mammalian nuclear localization sequences and many cell penetrating peptides as molecular sleds. These basic protein motifs exhibit weak but physiologically relevant sequence-nonspecific DNA affinity. Our findings indicate that many mammalian proteins contain molecular sled sequences and suggest the possibility that substantial undiscovered sliding activity exists among nuclear mammalian proteins. 相似文献
9.
组蛋白翻译后修饰是细胞DNA损伤早期应答反应的重要内涵,一方面是松弛、开放染色质结构的必要分子调节事件,以便DNA损伤响应蛋白能接近DNA损伤位点;另一方面直接参与DNA损伤修复蛋白招募过程的调控。综述了在DNA损伤信号激发下,发生的组蛋白主要修饰类型,异组蛋白H2AX、H2A.Z在DNA损伤部位与组蛋白置换,及其对DNA损伤响应蛋白招募的调节作用和机制。 相似文献
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《Critical reviews in biochemistry and molecular biology》2013,48(1-2):129-155
AbstractThe past decade has witnessed an exciting evolution in our understanding of eukaryotic DNA replication at the molecular level. Progress has been particularly rapid within the last few years due to the convergence of research on a variety of cell types, from yeast to human, encompassing disciplines ranging from clinical immunology to the molecular biology of viruses. New eukaryotic DNA replicases and accessory proteins have been purified and characterized, and some have been cloned and sequenced. In vitro systems for the replication of viral DNA have been developed, allowing the identification and purification of several mammalian replication proteins. In this review we focus on DNA polymerases alpha and delta and the polymerase accessory proteins, their physical and functional properties, as well as their roles in eukaryotic DNA replication. 相似文献
12.
Rosalie P. C. Driessen He Meng Gorle Suresh Rajesh Shahapure Giovanni Lanzani U Deva Priyakumar Malcolm F. White Helmut Schiessel John van Noort Remus Th. Dame 《Nucleic acids research》2013,41(1):196-205
Archaeal chromatin proteins share molecular and functional similarities with both bacterial and eukaryotic chromatin proteins. These proteins play an important role in functionally organizing the genomic DNA into a compact nucleoid. Cren7 and Sul7 are two crenarchaeal nucleoid-associated proteins, which are structurally homologous, but not conserved at the sequence level. Co-crystal structures have shown that these two proteins induce a sharp bend on binding to DNA. In this study, we have investigated the architectural properties of these proteins using atomic force microscopy, molecular dynamics simulations and magnetic tweezers. We demonstrate that Cren7 and Sul7 both compact DNA molecules to a similar extent. Using a theoretical model, we quantify the number of individual proteins bound to the DNA as a function of protein concentration and show that forces up to 3.5 pN do not affect this binding. Moreover, we investigate the flexibility of the bending angle induced by Cren7 and Sul7 and show that the protein–DNA complexes differ in flexibility from analogous bacterial and eukaryotic DNA-bending proteins. 相似文献
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An efficient DNA strand break repair contributes to the radioresistance of Deinococcus radiodurans, which harbors the DNA repair pathways nearly identical to Escherichia coli. The molecular mechanisms of these proteins functioning in 2 diverse classes of bacteria seem to be different. The macromolecular interactions and formation of multiprotein complexes in vivo have gained significant importance in explaining the mechanism of the complex cellular processes. Here, we report the identification of a novel DNA metabolic protein complex from D. radiodurans. A similar complex has, however, not been found in E. coli. Mass spectrometric analysis showed the presence of a few known DNA repair proteins, molecular chaperones, and a large number of uncharacterized proteins from D. radiodurans R1. Biochemical and immunoblotting results indicated the presence of the protein promoting DNA repair A, DNA polymerase, Mg2+, and (or) Mn2+ -dependent 5'-->3' exonuclease activity along with protein kinase activity and phosphoproteins. DNA ligase activity was completely dependent upon the ATP requirement, as no ligase activity was seen in the presence of NAD as a cofactor. These results suggest the molecular interactions of the known DNA repair proteins with uncharacterized proteins in the macromolecular complex and the regulation of DNA degradation with the involvement of ATP and protein kinase functions. 相似文献
14.
《Biophysical journal》2020,118(12):3008-3018
Protein diffusion in lower-dimensional spaces is used for various cellular functions. For example, sliding on DNA is essential for proteins searching for their target sites, and protein diffusion on microtubules is important for proper cell division and neuronal development. On the one hand, these linear diffusion processes are mediated by long-range electrostatic interactions between positively charged proteins and negatively charged biopolymers and have similar characteristic diffusion coefficients. On the other hand, DNA and microtubules have different structural properties. Here, using computational approaches, we studied the mechanism of protein diffusion along DNA and microtubules by exploring the diffusion of both protein types on both biopolymers. We found that DNA-binding and microtubule-binding proteins can diffuse on each other’s substrates; however, the adopted diffusion mechanism depends on the molecular properties of the diffusing proteins and the biopolymers. On the protein side, only DNA-binding proteins can perform rotation-coupled diffusion along DNA, with this being due to their higher net charge and its spatial organization at the DNA recognition helix. By contrast, the lower net charge on microtubule-binding proteins enables them to diffuse more quickly than DNA-binding proteins on both biopolymers. On the biopolymer side, microtubules possess intrinsically disordered, negatively charged C-terminal tails that interact with microtubule-binding proteins, thus supporting their diffusion. Thus, although both DNA-binding and microtubule-binding proteins can diffuse on the negatively charged biopolymers, the unique molecular features of the biopolymers and of their natural substrates are essential for function. 相似文献
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Specific interactions of DNA with proteins are required for both the replication of deoxyribonucleic acid proper and its regulation. Genetic elements of bacteria, their extrachromosomal elements in particular, represent a suitable model system for studies of these processes at the molecular level. In addition to replication enzymes (DNA polymerases), a series of other protein factors (e.g. topoisomerases, DNA unwinding enzymes, and DNA binding proteins) are involved in the replication of the chromosomal, phage and plasmid DNA. Specific interactions of proteins with DNA are particularly important in the regulation of initiation of DNA synthesis. Association of DNAs with the cell membrane also plays an important role in their replication in bacteria. 相似文献
17.
The inhibitor of growth (ING) family of proteins is an evolutionarily conserved family, with members present from yeast to humans. The mammalian ING proteins are candidate tumor suppressor proteins and accordingly can cooperate with p53 to arrest proliferation and induce apoptosis. ING proteins are also reported to function in the promotion of cellular senescence, the regulation of DNA damage responses and the inhibition of angiogenesis. At the molecular level, ING proteins are thought to function as chromatin regulatory molecules, acting as co-factors for distinct histone and factor acetyl-transferase (H/FAT) and deacetylase (HDAC) enzyme complexes. Further, ING proteins interact with a number of additional proteins involved in the regulation of critical nuclear processes, such as gene expression and DNA replication, and also function as nuclear phosphoinositide (PtdInsP) receptors. Despite the increasing number of known molecular interacting partners for ING proteins, the specific biochemical action of mammalian ING proteins and its relationship to tumor suppression remain elusive. In this Prospect, we summarize the present understanding of the binding partners and physiologic roles of ING proteins and propose a general molecular paradigm for how ING proteins might function to prevent cancer. 相似文献
18.
The biochemical mechanism and developmental expression for the repair of alkylated DNA has been characterized from Drosophila. As in other organisms, the correction of O6-methylguanine in Drosophila was found to involve the transfer of a methyl group from DNA to a protein cysteine residue. Two methylated proteins with subunit molecular weights of 30 kDa and 19 kDa were identified following incubation with [3H]-methylated substrate DNA and denaturing polyacrylamide gel electrophoresis. Identical molecular weights were found for the unmethylated forms of protein through their reaction to an antibody prepared against the 19 kDa Escherichia coli methyltransferase. Both Drosophila proteins are serologically reactive in adult males and females and most of the other developmental stages tested, with embryos representing the possible exception. The Drosophila proteins do not appear to be induced by sublethal exposures to alkylating agent. 相似文献
19.
Mitochondrial DNA plays a crucial role in cellular homeostasis; however, the molecular mechanisms underlying mitochondrial DNA inheritance and propagation are only beginning to be understood. To ensure the distribution and propagation of the mitochondrial genome, mitochondrial DNA is packaged into macromolecular assemblies called nucleoids, composed of one or more copies of mitochondrial DNA and associated proteins. We review current research on the mitochondrial nucleoid, including nucleoid-associated proteins, nucleoid dynamics within the cell, potential mechanisms to ensure proper distribution of nucleoids, and the impact of nucleoid organization on mitochondrial dysfunction. The nucleoid is the molecular organizing unit of mitochondrial genetics, and is the site of interactions that ultimately determine the bioenergetic state of the cell as a whole. Current and future research will provide essential insights into the molecular and cellular interactions that cause bioenergetic crisis, and yield clues for therapeutic rescue of mitochondrial dysfunction. 相似文献
20.
Nucleic acid isothermal amplification technologies: a review 总被引:3,自引:0,他引:3