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Identification, cloning, nucleotide sequence and chromosomal map location of hns, the structural gene for Escherichia coli DNA-binding protein H-NS 总被引:10,自引:0,他引:10
Cynthia L. Pon Raffaele A. Calogero Claudio O. Gualerzi 《Molecular & general genetics : MGG》1988,212(2):199-202
Summary Beginning with a synthetic oligonucleotide probe derived from its amino acid sequence, we have identified, cloned and sequenced the hns gene encoding H-NS, an abundant Escherichia coli 15 kDa DNA-binding protein with a possible histone-like function. The amino acid sequence of the protein deduced from the nucleotide sequence is in full agreement with that determined for H-NS. By comparison of the restriction map of the cloned gene and of its neighboring regions with the physical map of E. coli K12 as well as by hybridization of the hns gene with restriction fragments derived from the total chromosome, we have located the hns gene oriented counterclockwise at 6.1 min on the E. coli chromosome, just before an IS30 insertion element. 相似文献
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To study the mechanism of gamma-ray-induced illegitimate recombination, we examined the formation of lambdabio transducing phage in Escherichia coli after gamma-ray irradiation. We show that gamma-ray irradiation enhances the formation of lambdabio transducing phage during prophage induction. Moreover, an hns mutation synergistically enhanced the incidence of lambda-ray-induced illegitimate recombination. Next we determined the sequences at the recombination junctions of the lambdabio transducing phages induced by gamma-ray irradiation. Most of the recombination sites coincided with known hotspots. Among them, hotspot I accounted for 67% and 77% of gamma-ray-induced lambdabio transducing phages in the wild type and the hns mutant, respectively. Therefore, the recombination sites appear to occur mostly at hotspot I or at other hotspots, but rarely at non-hotspot sites. These results suggest that types of DNA damage other than the double-strand breaks induced at random sites are mainly responsible for the introduction of the site-specific or region-specific DNA double strand breaks that lead to recombination at the hotspots. The results also showed that the recombination events took place between DNA sequences possessing short stretches of homology. H-NS protein, which binds to curved DNA, suppresses illegitimate recombination in the presence and absence of gamma-ray irradiation. Models for gamma-ray-induced illegitimate recombination are discussed. 相似文献
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摘要:【目的】利用大肠杆菌BL21λDE3的表达系统,表达出有活性的鼠疫耶尔森氏菌(以下简称鼠疫菌)调控子蛋白H-NS,为进一步研究H-NS的转录调控奠定基础。【方法】 PCR扩增鼠疫菌201株hns基因的编码区,将其直接克隆入pET28a质粒中,再将pET28a-hns重组质粒转入大肠杆菌BL21λDE3菌株中,所得菌株经IPTG诱导后能表达出鼠疫菌His-H-NS蛋白;通过体外的凝胶迁移实验(EMSA)和DNaseⅠ足迹实验对His-H-NS蛋白与DNA的结合活性进行分析。【结果】成功表达出有活性的鼠疫菌His-H-NS蛋白,该蛋白对鼠疫菌pH6抗原基因(psaA、psaE)及rovA基因均有结合活性。【结论】鼠疫菌His-H-NS具有DNA结合活性,说明H-NS能调控鼠疫菌基因的转录。 相似文献
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Mutations in bglY, the structural gene for the DNA-binding protein H1, affect expression of several Escherichia coli genes 总被引:8,自引:0,他引:8
The protein patterns of a bglY mutant and the isogenic wild-type strains were compared on 2-dimensional gel electrophoresis. The synthesis of at least 36 peptides was affected. This suggests a global but specific role on gene expression for the DNA-binding protein H1. 相似文献
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A fragment of the DNA-binding protein of adenovirus type 5 has been obtained by controlled chymotryptic digestion of the entire molecule. Partial sequence determination indicates that the fragment consists of amino acids 174-525. The fragment is biologically active as measured by its ability to substitute for the entire molecule in a reconstituted DNA replication system. Crystals have been obtained that show diffraction to 2 A. 相似文献
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The hns gene is a member of the cold-shock regulon, indicating that the nucleoid-associated, DNA-binding protein H-NS plays an important role in the adaptation of Escherichia coli to low temperatures. We show here that the ability to cope efficiently with a cold environment (12°C and 25°C) is strongly impaired in E. coli strains carrying hns mutations. Growth inhibition is much more pronounced in strains carrying the hns-206 allele (an ampicillin resistance cassette inserted after codon 37) than in those carrying the hns-205 mutation (a Tn10 insertion located in codon 93). A protein fragment (H-NS*) is synthesized in strains carrying the hns-205::Tn10 mutation, suggesting that this truncated polypeptide is partially functional in the cold adaptation process. Analysis of the growth properties of strains harbouring four different low-copy-number plasmid-encoded hns genes that result in the production of C-terminally truncated H-NS proteins supports this proposal. H-NS* proteins composed of 133, 117 or 94 amino-terminal amino acids partially complemented the severe cold-sensitive growth phenotype of the hns-206 mutant. In contrast, synthesis of a truncated H-NS protein with only 75 amino-terminal amino acids was insufficient to restore growth at low temperature. 相似文献
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Smyth CP Lundbäck T Renzoni D Siligardi G Beavil R Layton M Sidebotham JM Hinton JC Driscoll PC Higgins CF Ladbury JE 《Molecular microbiology》2000,36(4):962-972
H-NS is a major component of the bacterial nucleoid, involved in condensing and packaging DNA and modulating gene expression. The mechanism by which this is achieved remains unclear. Genetic data show that the biological properties of H-NS are influenced by its oligomerization properties. We have applied a variety of biophysical techniques to study the structural basis of oligomerization of the H-NS protein from Salmonella typhimurium. The N-terminal 89 amino acids are responsible for oligomerization. The first 64 residues form a trimer dominated by an alpha-helix, likely to be in coiled-coil conformation. Extending this polypeptide to 89 amino acids generated higher order, heterodisperse oligomers. Similarly, in the full-length protein no single, defined oligomeric state is adopted. The C-terminal 48 residues do not participate in oligomerization and form a monomeric, DNA-binding domain. These N- and C-terminal domains are joined via a flexible linker which enables them to function independently within the context of the full-length protein. This novel mode of oligomerization may account for the unusual binding properties of H-NS. 相似文献
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Histone-like nucleoid structuring protein (H-NS) is a modular protein that is associated with the bacterial nucleoid. We used chromatin immunoprecipitation to determine the binding sites of H-NS and RNA polymerase on the Salmonella enterica serovar Typhimurium chromosome. We found that H-NS does not bind to actively transcribed genes and does not co-localize with RNA polymerase. This shows that H-NS principally silences gene expression by restricting the access of RNA polymerase to the DNA. H-NS had previously been shown to preferentially bind to curved DNA in vitro. In fact, at the genomic level we discovered that the level of H-NS binding correlates better with the AT-content of DNA. This is likely to have evolutionary consequences because we show that H-NS binds to many Salmonella genes acquired by lateral gene transfer, and functions as a gene silencer. The removal of H-NS from the cell causes un-controlled expression of several Salmonella pathogenicity islands, and we demonstrate that this has deleterious consequences for bacterial fitness. Our discovery of this novel role for H-NS may have implications for the acquisition of foreign genes by enteric bacteria. 相似文献
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The hns gene is a member of the cold-shock regulon, indicating that the nucleoid-associated, DNA-binding protein H-NS plays an important role in the adaptation of Escherichia coli to low temperatures. We show here that the ability to cope efficiently with a cold environment (12°C and 25°C) is strongly impaired in E. coli strains carrying hns mutations. Growth inhibition is much more pronounced in strains carrying the hns-206 allele (an ampicillin resistance cassette inserted after codon 37) than in those carrying the hns-205 mutation (a Tn10 insertion located in codon 93). A protein fragment (H-NS*) is synthesized in strains carrying the hns-205::Tn10 mutation, suggesting that this truncated polypeptide is partially functional in the cold adaptation process. Analysis of the growth properties of strains harbouring four different low-copy-number plasmid-encoded hns genes that result in the production of C-terminally truncated H-NS proteins supports this proposal. H-NS* proteins composed of 133, 117 or 94 amino-terminal amino acids partially complemented the severe cold-sensitive growth phenotype of the hns-206 mutant. In contrast, synthesis of a truncated H-NS protein with only 75 amino-terminal amino acids was insufficient to restore growth at low temperature. 相似文献
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Sandalli Cemal Özgümüş Osman Birol Sevim Ali 《World journal of microbiology & biotechnology》2010,26(11):2099-2103
Tetracycline resistance has been used as the key determinant to monitor resistance genes in natural environments such as rivers,
lakes or seawater. The aim of this study was evaluate the frequency of tetA, tetB, tetC, tetD and tetE genes in 52 tetracycline-resistant Enterobacteriacea isolated from river water in the North East Black Sea Region of Turkey. In 52 tetracycline-resistant strains, resistance
was mediated by tetA in eight (15.3%) strains, tetB in ten (19.2%) strains and both tetA and tetB in one (1.9%) strain. No tetC, tetD or tetE-mediated resistance was detected. In conclusion, the river water may be considered as a reservoir for the antibiotic resistance
genes and the people living in this area may be under risk. To our knowledge, this is the first report for molecular characterization
of tetracycline resistance in Enterobacteriaceae of river water origin in Turkey. 相似文献
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DNA-binding properties of the major structural protein of simian virus 40. 总被引:10,自引:7,他引:3
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T Soussi 《Journal of virology》1986,59(3):740-742
We investigated whether the VP1 protein of simian virus 40 binds to DNA. In vitro DNA-binding experiments clearly indicate that VP1 bound strongly to double-stranded and single-stranded DNA, with a higher affinity for the latter; additional experiments show that VP1 did not bind to a specific sequence of simian virus 40 DNA. 相似文献