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Population characteristics of Irish Helicobacter pylori isolates:: a tRNA-associated locus

IRISH JOURNAL OF MEDICAL SCIENCE 2000年第2期169卷 137-140页

作者： Dundon, WG Marshall, DG O'Moráin, CA Smyth, CJ Univ Dublin Trinity Coll Moyne Inst Prevent Med Dept Microbiol Dublin 2 Ireland

Background A strain-variable transfer RNA-associated-locus (trl) was present in 50% of Irish Helicobacter pylori (H. pylori), isolates and did not correlate with the origin of the isolates. Aim To associate a particular genotype or phenotype to trl status in H. pylori by further screening the isolates from the original study for the presence of known genotypic and phenotypic characteristics. Methods Forty two clinical isolates were screened for the presence of the cagA, vacA, iceA1 acid vapD genes by Southern or DNA dot blot analysis. Western blot analysis was performed using antibodies to CagA, VacA, Lewis X (Le(x)) and Lewis Y (Le(y)). Plasmids were identified by the alkaline lysis method. Results The cagA gene was present in 29 (69%) of isolates screened and 21 (50%) produced the CagA protein. The vacA gene was detected in all of the isolates while VacA was expressed in 71.4%. The iceA1 and vapD loci were detected in 73.8% and 71.4% respectively. Le(x) was expressed in 42.9% and Le(y) in 38.1% of the isolates. Expression of both Lewis antigens was detected in 7.1% while in 30.9% neither antigen was detected. Plasmids were present in 47.6%. There was no association between the trl status of isolates and any of the above. There were no significant associations between the phenotypic and genotypic characteristics studied and peptic ulcer disease or non-ulcer dyspepsia. Conclusion The strain-variable tRNA-associated locus is independent of the vacA/VacA, cagA/CagA, Lewis X, Lewis Y, iceA1, vapD and plasmid status in the population of Irish H. pylori isolates studied.

关键词：细菌蛋白质类/遗传学细菌蛋白质类/分离和提纯基因型幽门螺杆菌/遗传学幽门螺杆菌/分离和提纯爱尔兰表型 RNA 细菌/遗传学 RNA 转移/遗传学 RNA 转移甘氨酸/遗传学 RNA 转移亮氨酸/遗传学人类

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Cloning of a new Bacillus thuringiensis cry1I-type crystal protein

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CURRENT MICROBIOLOGY 2000年第1期41卷 65-69页

作者： Choi, SK Shin, BS Kong, EM Rho, HM Park, SH KRIBB Genome Res Ctr Taejon 305600 South Korea KRIBB Lab Microbial & Bioproc Engn Taejon 305600 South Korea Seoul Natl Univ Dept Mol Biol Seoul 151742 South Korea Seoul Natl Univ Res Ctr Cell Differentiat Seoul 151742 South Korea

A new cvy1I-type gene, cry1Id1, was cloned from a B. thuringiensis isolate, and its nucleotide sequence was determined. The deduced amino acid sequence of Cry1Id1 is 89.7%, 87.2%, and 83.4% identical to the Cry1Ia, Cry1Ib, and Cry1Ic proteins, respectively. The upstream sequence of the cry1Id1 structural gene was not functional as promoter in B. subtilis. The Cry1Id1 protein, purified from recombinant E. coli cells, had a toxicity comparable to that of Cry1Ia against Plutella xylostella, but it was significantly less active than Cry1Ia against Bombyx mori. Cry1Id1 was not active against the coleopteran insect, Agelastica coerulea.

关键词：氨基酸序列芽孢杆菌苏云金/遗传学芽孢杆菌苏云金/代谢细菌蛋白质类/遗传学细菌蛋白质类/分离和提纯细菌毒素类/遗传学细菌毒素类/分离和提纯碱基序列甲虫蚕属克隆分子内毒素类/遗传学内毒素类/分离和提纯溶血素蛋白质类杀虫药/分离和提纯分子序列数据蛾蛋白质结构三级动物

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Expression, purification, crystallization and preliminary X-ray diffraction studies of bacterial and archaeal L4 ribosomal proteins

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ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY 2000年第5期56卷 645-647页

作者： Worbs, M Wahl, MC Max Planck Inst Biochem Abt Strukturforsch D-82152 Martinsried Germany

Ribosomal protein L4 is implicated in the peptidyltransferase activity of the ribosome and in certain bacteria it regulates the transcription and translation of the 11-gene S10 operon. The genes for the L4 ribosomal proteins from the hyperthermophilic bacterium Thermotoga maritima and the halophilic archaeon Haloarcula marismortui have been PCR amplified from genomic DNA and cloned under the control of a T7 promoter to generate overexpressing Escherichia coli strains. For both proteins, efficient purification procedures were developed to yield material suitable for crystallization trials. Crystals of T. maritima L4 were obtained in the orthorhombic space group P2(1)2(1)2(1), with one molecule per asymmetric unit, diffracting to 1.7 Angstrom resolution with synchrotron radiation. Crystals of H. marismortui L4 belonged to the trigonal space group P3(1)21 or P3(2)21 and diffracted to 3.2 Angstrom resolution with a rotating-anode source, presumably containing three molecules per asymmetric unit. The results demonstrate that for certain halophilic proteins the same purification and crystallization procedures can be employed as for conventional proteins.

关键词：古细菌蛋白质类/化学古细菌蛋白质类/遗传学古细菌蛋白质类/分离和提纯细菌蛋白质类/化学细菌蛋白质类/遗传学细菌蛋白质类/分离和提纯克隆分子结晶结晶学 X线/方法死海嗜盐古细菌/遗传学蛋白质构象重组蛋白质类/化学重组蛋白质类/分离和提纯核糖体蛋白质类/化学核糖体蛋白质类/遗传学核糖体蛋白质类/分离和提纯喜温海洋杆菌/遗传学

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Crystallization and preliminary X-ray analysis of a membrane-bound nitrite reductase from Desulfovibrio desulfuricans ATCC 27774

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ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY 2000年第2期56卷 215-217页

作者： Dias, JM Cunha, CA Teixeira, S Almeida, G Costa, C Lampreia, J Moura, JJG Moura, I Romao, MJ Univ Nova Lisboa Fac Ciencias & Tecnol Ctr Quim Fina & Biotecnol Dept Quim P-2825114 Monte De Caparica Portugal Inst Tecnol Quim & Biol P-2780 Oeiras Portugal

Nitrite reductase from the sulfate-reducing bacterium Desulfovibrio desulfuricans ATCC 27774 is a multihaem (type c) membrane-bound enzyme that catalyzes the dissimilatory conversion of nitrite to ammonia. Crystals of the oxidized form of this enzyme were obtained using PEG and CaCl2 as precipitants in the presence of 3-(decylmethylammonium)propane-1-sulfonate and belong to the space group P2(1)2(1)2(1), With unit-cell parameters a = 78.94, b = 104.59, c = 143.18 Angstrom. A complete data set to 2.30 Angstrom resolution was collected using synchrotron radiation at the ESRF. However, the crystals may diffract to beyond 1.7 Angstrom and high-resolution data will be collected in the near future.

关键词：细菌蛋白质类/化学细菌蛋白质类/分离和提纯结晶脱硫弧菌属/分类脱硫弧菌属/酶学电泳聚丙烯酰氨凝胶膜蛋白质类/化学膜蛋白质类/分离和提纯亚硝酸盐还原酶类/化学亚硝酸盐还原酶类/分离和提纯氧化还原十二烷硫酸钠 X线衍射

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Crystallization and preliminary crystallographic analysis of the high-potential iron-sulfur protein from Thermochromatium tepidum

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ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY 2000年第5期56卷 656-658页

作者： Nogi, T Kobayashi, M Nozawa, T Miki, K Kyoto Univ Grad Sch Sci Dept Chem Sakyo Ku Kyoto 6068502 Japan Tohoku Univ Grad Sch Engn Dept Biomol Engn Aoba Ku Sendai Miyagi 9808579 Japan RIKEN Harima Inst SPring 8 Mikazuki Hyogo 6795148 Japan

The high-potential iron-sulfur protein (HiPIP) is an electron carrier between the photosynthetic reaction centre and the cytochrome bc(1) complex in the electron-transfer chain of photosynthesis. The purified HiPIP from Thermochromatium tepidum (formerly Chromatium tepidum) was crystallized in a solution of 1.4 M ammonium sulfate and 0.1 M sodium citrate pH 3.5. The crystals diffract X-rays beyond 1.4 Angstrom resolution and belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 47.12 (6), b = 59.59 (10), c = 23.62 (3) Angstrom. The structure was preliminarily solved by the molecular-replacement method. The crystal structure of HiPIP from T. tepidum showed that the proteins exist as monomers, although HiPIPs from several other species can form dimers.

关键词：细菌蛋白质类/化学细菌蛋白质类/分离和提纯色菌属/代谢结晶结晶学 X线/方法二聚作用铁硫蛋白质类/化学铁硫蛋白质类/分离和提纯光合作用中心复合蛋白质类

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Crystallization and preliminary diffraction studies of a truncated form of a novel protease from spores of Bacillus megaterium

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ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY 2000年第1期56卷 70-72页

作者： Ponnuraj, K Kelly, S Nessi, C Setlow, P Jedrzejas, MJ Univ Alabama Birmingham Dept Microbiol Birmingham AL 35294 USA Univ Connecticut Ctr Hlth Dept Biochem Farmington CT 06030 USA

During germination of spores of Bacillus species, a novel protease termed GPR initiates the degradation of a group of small acid-soluble spore proteins which protect the dormant spore's DNA from damage. Trypsin digestion of the zymogen of B. megaterium GPR removes similar to 15 kDa from the C-terminal end of the 46 kDa zymogen subunit, leaving a 30 kDa subunit. Single crystals of this truncated form of GPR have been obtained by the vapor-diffusion method using PEG 4000 as a precipitating agent. The crystals belong to the monoclinic space group P2(1), with unit-cell parameters a = 67.99, b = 105.34, c = 108.63 Angstrom, beta = 95.68 degrees. The cryofrozen crystals diffract X-rays to about 3.3 Angstrom using synchrotron radiation.

关键词：巨大芽孢杆菌/酶学细菌蛋白质类/化学细菌蛋白质类/分离和提纯结晶结晶学 X线内肽酶类/化学内肽酶类/分离和提纯分子量 ∑因子孢子细菌/酶学转录因子胰蛋白酶

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Purification, crystallization and preliminary X-ray crystallographic study of α-amylase from Bacillus stearothermophilus

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ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY 2000年第Pt 2期56卷 200-202页

作者： Suvd, D Takase, K Fujimoto, Z Matsumura, M Mizuno, H Univ Tsukuba Inst Appl Biochem Tsukuba Ibaraki 3058572 Japan Natl Inst Agrobiol Resources Dept Biotechnol Tsukuba Ibaraki 3058602 Japan

A recombinant alpha-amylase from Bacillus stearothermophilus was found to be produced as several isoforms arising from different N-terminal processing. Some of those isoforms were purified to homogeneity and crystallized at 293 K using the hanging-drop vapour-diffusion method under the following conditions: 35 mM sodium acetate (pH 4.6), 6.25% (nu/nu) 2-propanol, in the presence of 1.23% (w/nu) acarbose (a pseudo-oligosaccharide inhibitor) in the drop. The crystals diffracted beyond 2.0 Angstrom resolution using synchrotron radiation at the Photon Factory, Tsukuba. They belong to the monoclinic space group P2(1), with unit-cell parameters a = 53.7 (2), b = 92.9 (4), c = 53.2 (2) Angstrom, beta = 109.4 (1)degrees.

关键词：氨基酸序列细菌蛋白质类/化学细菌蛋白质类/分离和提纯结晶结晶学 X线嗜热脂肪芽孢杆菌/酶学分子序列数据蛋白质亚型/化学蛋白质亚型/分离和提纯重组蛋白质类/化学重组蛋白质类/分离和提纯 α淀粉酶类/化学 α淀粉酶类/分离和提纯

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Identification of a lexA gene in, and construction of a lexA mutant of, Xanthomonas campestris pv. citri

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CURRENT MICROBIOLOGY 2000年第4期40卷 233-238页

作者： Yang, MK Wu, PI Yang, YC Fu Jen Catholic Univ Dept Biol Taipei 242 Taiwan

The lexA gene of Xanthomonas campestris pathovar citri (X.c. pv. citri) was cloned and sequenced. The 639-bp open reading frame encodes a protein of 213 amino acids that shares substantial sequence homology with the products of previously characterized lexA genes, sharing 46% identity with the LexA protein of Escherichia coli. Amino acids required for autocatalytic cleavage of LexA are conserved in the X.c. pv. citri protein, whereas domains thought to mediate DNA binding differ markedly from those of LexA proteins from E. coli and other bacteria. The X.c. pv. citri LexA protein was overexpressed in E. coli, and SDS-polyacrylamide gel electrophoresis revealed a molecular size of 23 kDa for the purified protein. A lexA mutant of X.c. pv. citri was constructed by gene replacement, and the basal level of recA expression in this mutant was shown to be similar to that for wild-type cells exposed to a DNA-damaging agent. These results indicate that LexA functions as a repressor of recA expression in X.c. pv. citri.

关键词：细菌蛋白质类/遗传学细菌蛋白质类/分离和提纯细菌蛋白质类/药理学碱基序列印迹法 DNA 克隆分子 DNA 细菌/分析电泳聚丙烯酰氨凝胶大肠杆菌/遗传学基因表达调控/药物作用基因细菌分子序列数据突变 RecA重组酶/遗传学重组蛋白质类/遗传学序列同源性氨基酸序列同源性核酸丝氨酸内肽酶类/遗传学丝氨酸内肽酶类/分离和提纯丝氨酸内肽酶类/药理学黄单胞菌野油菜/化学黄单胞菌野油菜/遗传学

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Crystallization and preliminary X-ray analyses of catabolite control protein A, free and in complex with its DNA-binding site

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ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY 2000年第Pt 1期56卷 67-69页

作者： Tebbe, J Orth, P Küster-Schöck, E Hillen, W Saenger, W Hinrichs, W Free Univ Berlin Inst Kristallog D-14195 Berlin Germany Univ Erlangen Nurnberg Lehrstuhl Mikrobiol Inst Mikrobiol Biochem & Genet D-91058 Erlangen Germany

The catabolite control protein (CcpA) from Bacillus megaterium is a member of the bacterial repressor protein family GalR/LacI. CcpA with an N-terminal His-tag was used for crystallization. Crystals of free CcpA and of CcpA in complex with the putative operator sequence (catabolite responsive elements, CRE) were obtained by vapour-diffusion techniques at 291 K using the hanging-drop method. CcpA crystals grown in the presence of polyethylene glycol 8000 belong to the hexagonal space group P6(1)22 or P6(5)22, with unit-cell parameters a = 74.4, c = 238.8 Angstrom. These crystals diffract X-rays to 2.55 Angstrom resolution and contain one monomer of the homodimeric protein per asymmetric unit. Crystals of the CcpA-CRE complex were obtained with ammonium sulfate as precipitant and belong to the tetragonal space group I4(1)22, with unit-cell parameters a = 125, c = 400 Angstrom and one complex per asymmetric unit. Although these cocrystals grew to a sufficient size, X-ray diffraction was limited to 8 Angstrom resolution.

关键词：氨基酸序列巨大芽孢杆菌/化学巨大芽孢杆菌/遗传学细菌蛋白质类/化学细菌蛋白质类/遗传学细菌蛋白质类/分离和提纯碱基序列结合部位结晶结晶学 X线 DNA 细菌/化学 DNA 细菌/遗传学 DNA 细菌/分离和提纯 DNA结合蛋白质类/化学 DNA结合蛋白质类/遗传学 DNA结合蛋白质类/分离和提纯大肠杆菌/遗传学重组蛋白质类/化学重组蛋白质类/遗传学重组蛋白质类/分离和提纯阻遏蛋白质类/化学阻遏蛋白质类/遗传学阻遏蛋白质类/分离和提纯

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Characterisation of the catalytically active form of RecG helicase

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NUCLEIC ACIDS RESEARCH 2000年第12期28卷 2324-2332页

作者： McGlynn, P Mahdi, AA Lloyd, RG Univ Nottingham Queens Med Ctr Inst Genet Nottingham NG7 2UH England

Replication of DNA is fraught with difficulty and chromosomes contain many lesions which may block movement of the replicative machinery. However, several mechanisms to overcome such problems are beginning to emerge from studies with Escherichia coli. An important enzyme in one or more of these mechanisms is the RecG helicase, which may target stalled replication forks to generate a four-stranded (Holliday) junction, thus facilitating repair and/or bypass of the original lesion. To begin to understand how RecG might catalyse regression of fork structures, we have analysed what the catalytically active form of the enzyme may be. We have found that RecG exists as a monomer in solution as measured by gel filtration but when bound to junction DNA the enzyme forms two distinct protein-DNA complexes that contain one and two protein molecules, However, mutant inhibition studies failed to provide any evidence that RecG acts as a multimer in vitro. Additionally, there was no evidence for cooperativity in the junction DNA-stimulated hydrolysis of ATP. These data suggest that RecG functions as a monomer to unwind junction DNA, which supports an 'inchworm' rather than an 'active rolling' mechanism of DNA unwinding. The observed in vivo inhibition of wild-type RecG by mutant forms of the enzyme was attributed to occlusion of the DNA target and correlates with the very low abundance of replication forks within an *** cell, even during rapid growth.

关键词：氨基酸序列氨基酸取代细菌蛋白质类/遗传学细菌蛋白质类/分离和提纯细菌蛋白质类/代谢碱基序列结合部位催化作用色谱法亲和 DNA解旋酶类/代谢大肠杆菌/酶学大肠杆菌蛋白质类动力学分子序列数据诱变定点寡脱氧核糖核苷酸类/化学寡脱氧核糖核苷酸类/代谢重组融合蛋白质类/化学重组融合蛋白质类/代谢底物特异性模板遗传

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