Journal of Microbiology

Volume 50(1); February 2012

Review

REVIEW] Recent Findings about the Yersinia enterocolitica Phage Shock Protein Response: Saori Yamaguchi , Andrew J. Darwin; J. Microbiol. 2012;50(1):1-7. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1578-7

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Abstract: The phage shock protein (Psp) system is a conserved extracytoplasmic stress response in bacteria that is essential for virulence of the human pathogen Yersinia enterocolitica. This article summarizes some recent findings about Y. enterocolitica Psp system function. Increased psp gene expression requires the transcription factor PspF, but under non-inducing conditions PspF is inhibited by an interaction with another protein, PspA, in the cytoplasm. A Psp-inducing stimulus causes PspA to relocate to the cytoplasmic membrane, freeing PspF to induce psp gene expression. This PspA relocation requires the integral cytoplasmic membrane proteins, PspB and PspC, which might sense an inducing trigger and sequester PspA by direct interaction. The subsequent induction of psp gene expression increases the PspA concentration, which also allows it to contact the membrane directly, perhaps for its physiological function. Mutational analysis of the PspB and PspC proteins has revealed that they both positively and negatively regulate psp gene expression and has also identified PspC domains associated with each function. We also compare the contrasting physiological roles of the Psp system in the virulence of Y. enterocolitica and Salmonella enterica sv. Typhimurium (S. Typhimurium). In S. Typhimurium, PspA maintains the proton motive force, which provides the energy needed to drive ion importers required for survival within macrophages. In contrast, in the extracellular pathogen Y. enterocolitica, PspB and PspC, but not PspA, are the Psp components needed for virulence. PspBC protect Y. enterocolitica from damage caused by the secretin component of its type 3 secretion system, an essential virulence factor.

Research Support, Non-U.S. Gov'ts

Stratified Distribution of Nutrients and Extremophile Biota within Freshwater Ice Covering the Surface of Lake Baikal: Nina A. Bondarenko , Olga I. Belykh , Ludmila P. Golobokova , Olga V. Artemyeva , Natalia F. Logacheva , Irina V. Tikhonova , Irina A. Lipko , Tatyana Ya. Kostornova , Valentina V. Parfenova , Tamara V. Khodzher , Young-Gun Zo; J. Microbiol. 2012;50(1):8-16. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1251-1

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Abstract: Biological entities and gradients of selected chemicals within the seemingly barren ice layers covering Lake Baikal were investigated. Ice cores 40–68 cm long were obtained from inshore and offshore sites of Southern Lake Baikal during the cold period of a year (March-April) in 2007 and 2008. In microscopic observations of the melted ice, both algae and bacteria were found in considerable numbers (>103 cells/L and >104 cells/ml, respectively). Among all organisms found, diatom was generally the most predominant taxon in the ice. Interestingly, both planktonic and benthic algae were present in considerable numbers (2–4×104 cells/L). Dominant phototrophic picoplankton were comprised of small green algae of various taxa and cyanobacteria of Synechococcus and Cyanobium. The bacterial community consisted mostly of short rod and cocci cells, either freeliving or aggregated. Large numbers of yeast-like cells and actinomycete mycelium were also observed. Concentrations of silica, phosphorus, and nitrate were low by an order of magnitude where biota was abundant. The profile of the ice could be interpreted as vertical stratification of nutrients and biomass due to biological activities. Therefore, the organisms in the ice were regarded to maintain high activity while thriving under freezing conditions. Based on the results, it was concluded that the freshwater ice covering the surface of Lake Baikal is considerably populated by extremophilic microorganisms that actively metabolize and form a detritus food chain in the unique large freshwater ecosystem of Lake Baikal.

Effects of Phosphate Addition on Biofilm Bacterial Communities and Water Quality in Annular Reactors Equipped with Stainless Steel and Ductile Cast Iron Pipes: Hyun-Jung Jang , Young-June Choi , Hee-Myong Ro , Jong-Ok Ka; J. Microbiol. 2012;50(1):17-28. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1040-x

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Abstract: The impact of orthophosphate addition on biofilm formation and water quality was studied in corrosion-resistant stainless steel (STS) pipe and corrosion-susceptible ductile cast iron (DCI) pipe using cultivation and culture-independent approaches. Sample coupons of DCI pipe and STS pipe were installed in annular reactors, which were operated for 9 months under hydraulic conditions similar to a domestic plumbing system. Addition of 5 mg/L of phosphate to the plumbing systems, under low residual chlorine conditions, promoted a more significant growth of biofilm and led to a greater rate reduction of disinfection by-products in DCI pipe than in STS pipe. While the level of THMs (trihalomethanes) increased under conditions of low biofilm concentration, the levels of HAAs (halo acetic acids) and CH (chloral hydrate) decreased in all cases in proportion to the amount of biofilm. It was also observed that chloroform, the main species of THM, was not readily decomposed biologically and decomposition was not proportional to the biofilm concentration; however, it was easily biodegraded after the addition of phosphate. Analysis of the 16S rDNA sequences of 102 biofilm isolates revealed that Proteobacteria (50%) was the most frequently detected phylum, followed by Firmicutes (10%) and Actinobacteria (2%), with 37% of the bacteria unclassified. Bradyrhizobium was the dominant genus on corroded DCI pipe, while Sphingomonas was predominant on non-corroded STS pipe. Methylobacterium and Afipia were detected only in the reactor without added phosphate. PCR-DGGE analysis showed that the diversity of species in biofilm tended to increase when phosphate was added regardless of the pipe material, indicating that phosphate addition upset the biological stability in the plumbing systems.

Host Species as a Strong Determinant of the Intestinal Microbiota of Fish Larvae: Xuemei Li , Yuhe Yu , Weisong Feng , Qingyun Yan , Yingchun Gong; J. Microbiol. 2012;50(1):29-37. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1340-1

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Abstract

We investigated the influence of host species on intestinal microbiota by comparing the gut bacterial community structure of four cohabitating freshwater fish larvae, silver carp, grass carp, bighead carp, and blunt snout bream, using denaturing gradient gel electrophoresis (DGGE) of the amplified 16S and 18S rRNA genes. Similarity clustering indicated that the intestinal microbiota derived from these four fish species could be divided into four groups based on 16S rRNA gene similarity, whereas the eukaryotic 18S rRNA genes showed no distinct groups. The water sample from the shared environment contained microbiota of an independent group as indicated by both 16S and 18S rRNA genes segments. The bacterial community structures were visualized using rank-abundance plots fitted with linear regression models.
Results
showed that the intestinal bacterial evenness was significantly different between species (P<0.05) and between species and the water sample (P<0.01). Thirty-five relatively dominant bands in DGGE patterns were sequenced and grouped into five major taxa: Proteobacteria (26), Actinobacteria (5), Bacteroidetes (1), Firmicutes (2), and Cyanobacterial (1). Six eukaryotes were detected by sequencing 18S rRNA genes segments. The present study suggests that the intestines of the four fish larvae, although reared in the same environment, contained distinct bacterial populations, while intestinal eukaryotic microorganisms were almost identical.

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Gram-positive Rhizobacterium Bacillus amyloliquefaciens FZB42 Colonizes Three Types of Plants in Different Patterns: Ben Fan , Rainer Borriss , Wilfrid Bleiss , Xiaoqin Wu; J. Microbiol. 2012;50(1):38-44. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1439-4

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Abstract: The colonization of three types of different plants, Zea mays, Arabidopsis thaliana, and Lemna minor, by GFP-labeled Gram-positive rhizobacterium Bacillus amyloliquefaciens FZB42 was studied in gnotobiotic systems using confocal laser scanning microscopy and electron microscopy. It was demonstrated that FZB42 was able to colonize all the plants. On one hand, similar to some Gram-negative rhizobacteria like Pseudomonas, FZB42 favored the areas such as the concavities in root surfaces and the junctions where lateral roots occurred from the primary roots; on the other hand, we clearly demonstrated that root hairs were a popular habitat to the Gram-positive rhizobacterium. FZB42 exhibited a specific colonization pattern on each of the three types of plants. On Arabidopsis, tips of primary roots were favored by FZB42 but not so on maize. On Lemna, FZB42 accumulated preferably along the grooves between epidermal cells of roots and in the concave spaces on ventral sides of fronds. The results suggested L. minor to be a promising tool for investigations on plant-microbial interaction due to a series of advantages it has. Colonization of maize and Arabidopsis roots by FZB42 was also studied in the soil system. Comparatively, higher amount of FZB42 inoculum (~108 CFU/ml) was required for detectable root colonization in the soil system, where the preference of FZB42 cells to root hairs were also observed.

Growth Promotion of Xanthium italicum by Application of Rhizobacterial Isolates of Bacillus aryabhattai in Microcosm Soil: Sol Lee , Jong-Ok Ka , Hong-Gyu Song; J. Microbiol. 2012;50(1):45-49. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1415-z

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Abstract

This study was conducted using rhizobacteria, which are able to exert beneficial effects upon plant growth in the infertile soil collected from barren lakeside areas. Four strains of plant growth promoting bacteria were isolated from the rhizosphere of a common wild plant, Erigeron canadensis. Isolated strains LS9, LS11, LS12, and LS15 were identified as Bacillus aryabhattai by 16S rDNA sequence analysis. B. aryabhattai LS9, LS11, LS12, and LS15 could solubilize 577.9, 676.8, 623.6, and 581.3 mg/L of 0.5% insoluble calcium phosphate within 2 days of incubation. Production of indole acetic acid, a typical growth promoting phytohormone auxin, by strain LS15 was 471.3 mg/L in 2 days with the addition of auxin precursor L-tryptophan. All the strains also produced other phytohormones such as indole butyric acid, gibberellins, and abscisic acid, and strain LS15 showed the highest production rate of gibberellin (GA3), 119.0 μg/mg protein. Isolated bacteria were used in a microcosm test for growth of wild plant Xanthium italicum, which can be utilized as a pioneer plant in barren lands. Seed germination was facilitated, and the lengths of roots, and shoots and the dry weights of germinated seedlings after 16 days were higher than those of the uninoculated control plants. Root lengths of seedlings of X. italicum increased by 121.1% in LS11-treated samples after 16 days. This plant growth-promoting capability of B. aryabhattai strains may be utilized as an environmentally friendly means of revegetating barren lands, especially sensitive areas such as lakeside lands.

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Diversity and Physiological Properties of Root Endophytic Actinobacteria in Native Herbaceous Plants of Korea: Tae-Ui Kim , Sung-Heun Cho , Ji-Hye Han , Young Min Shin , Hyang Burm Lee , Seung Bum Kim; J. Microbiol. 2012;50(1):50-57. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1417-x

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Abstract

Endophytic actinobacterial diversity in the native herbaceous plant species of Korea was analyzed using a culturebased approach. Sixty one actinobacterial strains were isolated, and assigned to 15 genera based on 16S rRNA gene analysis. The members of the genus Streptomyces comprised 45.9% of the total isolates, followed by Micromonospora (18.8%), Rhodococcus (6.6%), Microbispora (4.9%), and Micrococcus (4.9%). Other minor constituents included members of Microbacterium, Streptacidiphilus, Arthrobacter, Dietzia, Kitasatospora, Herbiconiux, Mycobacterium, Nocardia, Rathayibacter, and Tsukamurella. Among the isolates, 65.6% exhibited at least one hydrolytic enzyme activity out of four, and 45.9% exhibited antagonistic activity against at least one fungal pathogen out of five, thus demonstrating that endophytic actinobacteria can be an important source of bioactive compounds. Notably, most strains of Streptomyces proved active for both enzymatic and antagonistic activities.

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Microflora Profiling of Infected Root Canal before and after Treatment Using Culture-Independent Methods: Yasuhiro Ito , Takuichi Sato , Keiko Yamaki , Gen Mayanagi , Kazuhiro Hashimoto , Hidetoshi Shimauchi , Nobuhiro Takahashi; J. Microbiol. 2012;50(1):58-62. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-0459-4

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Abstract: This study aimed to profile the microflora in infected root canals before and after root canal treatment using cultureindependent
methods
. Six infected root canals in singlerooted teeth with periapical lesions from five subjects were included. Quantification of total bacteria was performed by real-time PCR with primers targeting 16S rRNA genes. PCR products with universal 16S rRNA gene primers were cloned and partially sequenced, and bacterial identification at the species level was performed by comparative analysis with the GenBank database. The concentration of extracted DNA before treatment was higher than that after root canal treatment, although the difference was not statistically significant. Sequence analysis revealed that oral bacteria such as Fusobacterium, Streptococcus, Olsenella, and Pseudoramibacter detected in cases before root canal treatment disappeared after treatment. These results suggest that the root canal microflora are distinct before and after root canal treatment, and that treatment changes the microflora in both quantity and quality.

Phenotypic and Phylogenetic Analysis of Lactic Acid Bacteria Isolated from Forage Crops and Grasses in the Tibetan Plateau: Huili Pang , Zhongfang Tan , Guangyong Qin , Yanping Wang , Zongwei Li , Qingsheng Jin , Yimin Cai; J. Microbiol. 2012;50(1):63-71. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1284-5

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Abstract

A total of 140 lactic acid bacteria (LAB) strains were isolated from corn, alfalfa, clover, sainfoin, and Indian goosegrass in the Tibetan Plateau. According to phenotypic and chemotaxonomic characteristics, 16S rDNA sequence, and recA gene PCR amplification, these LAB isolates were identified as belonging to five genera and nine species. Corn contained more LAB species than other forage crops. Leuconostoc pseudomesenteroides, Lactococcus lactis subsp. lactis, Lactobacillus brevis, and Weissella paramesenteroides were dominant members of the LAB population on alfalfa, clover, sainfoin, and Indian goosegrass, respectively. The comprehensive 16S rDNA and recA-based approach effectively described the LAB community structure of the relatively abundant LAB species distributed on different forage crops. This is the first report describing the diversity and natural populations of LAB associated with Tibetan forage crops, and most isolates grow well at or below 10°C. The results will be valuable for the future design of appropriate inoculants for silage fermentation in this very cold area.

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Journal Article

Purification and Characterization of a Novel Laccase from the Edible Mushroom Hericium coralloides: Ya-Jie Zou , He-Xiang Wang , Tzi-Bun Ng , Chen-Yang Huang , Jin-Xia Zhang; J. Microbiol. 2012;50(1):72-78. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1372-6

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Abstract: A novel laccase from the edible mushroom Hericium coralloides was purified by ion exchange chromatography on diethylaminoethyl (DEAE) cellulose, carboxymethyl (CM) cellulose, and Q-Sepharose columns followed by fast protein liquid chromatography gel filtration on a Superdex 75 column. Analysis by gel filtration and SDS-PAGE indicated that the protein is a monomer in solution with a molecular mass of 65 kDa. Its N-terminal amino acid sequence was AVGDDTPQLY, which exhibits partial sequence homology to previously isolated laccases. Optimum activity was observed at pH 2.2 and at 40°C. The enzyme showed activity toward a variety of substrates, the most sensitive of which was 2,2􍿁-azinobis [3-ethylbenzothiazolone-6-sulfonic acid] diammonium salt (ABTS). The degradation activity toward substrates was ABTS > N,N-dimethyl-1,4-phenylenediamine > catechol > 2-methylcatechol > pyrogallol. The laccase did not exert any antiproliferative activity against Hep G2 or MCF 7 tumor cell lines at a concentration of 60 μM, unlike some previously reported mushroom proteins, but showed significant activity toward human immunodeficiency virus-1 (HIV-1) reverse transcriptase with an IC50 of 0.06 μM.

Research Support, Non-U.S. Gov'ts

The PseEF Efflux System Is a Virulence Factor of Pseudomonas syringae pv. syringae: Hyosun Cho , Hyojeung Kang; J. Microbiol. 2012;50(1):79-90. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1353-9

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Abstract

An ATP-binding cassette (ABC) transporter, called the PseEF efflux system, was identified at the left border of the syr-syp genomic island of Pseudomonas syringae pv. syringae strain B301D. The PseEF efflux system was located within a 3.3-kb operon that encodes a periplasmic membrane fusion protein (PseE), and an ABC-type cytoplasmic membrane protein (PseF). The PseEF efflux system exhibited amino acid homology to a putative ABC efflux system (MacAB) of E. coli W3104 with identities of 47.2% (i.e., PseE to MacA) and 57.6% (i.e., PseF to MacB). A nonpolar mutation within the pseF gene was generated by nptII insertional mutagenesis. The resultant mutant strain showed significant reduction in secretion of syringomycin (74%) and syringopeptin (71%), as compared to parental strain B301D. Quantitative real-time RT-PCR was used to determine transcript levels of the syringomycin (syrB1) and syringopeptin (sypA) synthetase genes in strain B301D-HK7 (a pseF mutant). Expression of the sypA gene by mutant strain B301D-HK7 was approximately 6.9% as compared to that of parental strain B301D, while the syrB1 gene expression by mutant strain B301D-HK7 was nearly 14.6%. In addition, mutant strain B301D-HK7 was less virulent by approximately 67% than parental strain B301D in immature cherry fruits. Mutant strain B301D-HK7 was not reduced in resistance to any antibiotics used in this study as compared to parental strain B301D. Expression (transcript levels) of the pseF gene was induced approximately six times by strain B301D grown on syringomycin minimum medium (SRM) supplemented with the plant signal molecules arbutin and D-fructose (SRMAF), as compared to that of strain B301D grown on SRM (in the absence of plant signal molecules). In addition, during infection of bean plants by P. syringae pv. syringae strain B728a, expression of the pseF gene increased at 3 days after inoculation (dai). More than 180-fold induction was observed in transcript levels of the pseF gene by parental strain B728a as compared to strain B728a-SL7 (a salA mutant). Thus, the PseEF efflux system, an ABC-type efflux system, has an important role in secretion of syringomycin and syringopeptin, and is required for full virulence in P. syringae pv. syringae.

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Effects of a Dark-Septate Endophytic Isolate LBF-2 on the Medicinal Plant Lycium barbarum L.: Hai-han Zhang , Ming Tang , Hui Chen , Ya-jun Wang; J. Microbiol. 2012;50(1):91-96. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1159-9

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Abstract

Dark septate endophytes (DSE) are ubiquitous root associated fungi; however, our understanding of their ecological function remains unclear. Here, we investigated the positive effect of a DSE fungus on its host plant Lycium barbarum L. A DSE isolate, LBF-2, isolated from the roots of L. barbarum, was inoculated onto the roots of plants, which were grown under greenhouse conditions for five weeks. The result of molecular analyses of internal transcribed spacer regions indicated that LBF-2 was 96% similar to Paraphoma chrysanthemicola. Melanized septate hyphae were observed in the root cortical cells of L. barbarum using a light microscope. Inoculation with LBF-2 increased the total biomass by 39.2% and also enhanced chlorophyll fluorescence. Inoculation increased the concentration of total chlorophyll by 22.8% and of chlorophyll a by 21.3%, relative to uninoculated controls. These data indicate that the LBF-2 isolate might be used to facilitate the cultivation of L. barbarum, which has medicinal applications.

Citations

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Heat- and Cold-Shock Responses in Fusarium graminearum 3 Acetyl- and 15 Acetyl-Deoxynivalenol Chemotypes: Vladimir Vujanovic , Yit Kheng Goh , Prasad Daida; J. Microbiol. 2012;50(1):97-102. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1381-5

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Abstract: Fusarium graminearum Schwabe is the primary cause of Fusarium head blight (FHB) in North America. Chemically distinct F. graminearum sub-populations can be identified based on the type or composition of deoxynivalenol (DON) mycotoxin derivatives, including 3-acetyl (3-ADON) and 15-acetyl (15-ADON). The evaluation of randomly selected 3-ADON and 15-ADON isolates, collected from spring wheat throughout Canada, was performed using thin layer chromatography (TLC), high-performance liquid chromatography (HPLC), ice-nucleation activity (INA), and heat and cold tolerance tests conducted within a temperature range of -70°C to 65°C. The results indicated that the 3-ADON sub-population, which is responsible for the highest disease severity and has rapidly displaced the 15-ADON sub-population, produces more DON and zearalenone (ZEA) than the 15-ADON sub-population when exposed to heat and cold. Following exposures (1 and 2 h) to extremely high or low temperatures, 3-ADON isolates exhibited faster mycelial growth than 15-ADON isolates. In addition, the warmest temperature at which INA activity occurred was in 3-ADON (-3.6°C) vs. 15-ADON (-5.1°C). Taken together, these features suggest that the newly emerging 3-ADON sub-population is more resilient than the resident 15-ADON sub-population. Overall, the differences between the two sub-populations could provide new insights into FHB epidemiology and if validated under field conditions, may provide important information for predicting future FHB epidemics.

Journal Article

Chitinase Production by Bacillus thuringiensis and Bacillus licheniformis: Their Potential in Antifungal Biocontrol: Eman Zakaria Gomaa; J. Microbiol. 2012;50(1):103-111. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1343-y

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136 Scopus

Abstract: Thirty bacterial strains were isolated from the rhizosphere of plants collected from Egypt and screened for production of chitinase enzymes. Bacillus thuringiensis NM101-19 and Bacillus licheniformis NM120-17 had the highest chitinolytic activities amongst those investigated. The production of chitinase by B. thuringiensis and B. licheniformis was optimized using colloidal chitin medium amended with 1.5% colloidal chitin, with casein as a nitrogen source, at 30°C after five days of incubation. An enhancement of chitinase production by the two species was observed by addition of sugar substances and dried fungal mats to the colloidal chitin media. The optimal conditions for chitinase activity by B. thuringiensis and B. licheniformis were at 40°C, pH 7.0 and pH 8.0, respectively. Na+, Mg2+, Cu2+, and Ca2+ caused enhancement of enzyme activities whereas they were markedly inhibited by Zn2+, Hg2+, and Ag+. In vitro, B. thuringiensis and B. licheniformis chitinases had potential for cell wall lysis of many phytopathogenic fungi tested. The addition of B. thuringiensis chitinase was more effective than that of B. licheniformis in increasing the germination of soybean seeds infected with various phytopathogenic fungi.

Research Support, Non-U.S. Gov't

Saccharomyces cerevisiae Cmr1 Protein Preferentially Binds to UV-Damaged DNA In Vitro: Do-Hee Choi , Sung-Hun Kwon , Joon-Ho Kim , Sung-Ho Bae; J. Microbiol. 2012;50(1):112-118. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1597-4

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13 Scopus

Abstract: DNA metabolic processes such as DNA replication, recombination, and repair are fundamentally important for the maintenance of genome integrity and cell viability. Although a large number of proteins involved in these pathways have been extensively studied, many proteins still remain to be identified. In this study, we isolated DNA-binding proteins from Saccharomyces cerevisiae using DNA-cellulose columns. By analyzing the proteins using mass spectrometry, an uncharacterized protein, Cmr1/YDL156W, was identified. Cmr1 showed sequence homology to human Damaged-DNA binding protein 2 in its C-terminal WD40 repeats. Consistent with this finding, the purified recombinant Cmr1 protein was found to be intrinsically associated with DNA-binding activity and exhibited higher affinity to UV-damaged DNA substrates. Chromatin isolation experiments revealed that Cmr1 localized in both the chromatin and supernatant fractions, and the level of Cmr1 in the chromatin fraction increased when yeast cells were irradiated with UV. These
results
suggest that Cmr1 may be involved in DNA-damage responses in yeast.

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