• Journal of Microbiology and Biotechnology
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• v.27 no.2
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• pp.321-334
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• 2017

Process surveillance within agricultural biogas plants (BGPs) was concurrently studied by high-throughput 16S rRNA gene amplicon sequencing and an optimized quantitative microscopic fingerprinting (QMF) technique. In contrast to 16S rRNA gene amplicons, digitalized microscopy is a rapid and cost-effective method that facilitates enumeration and morphological differentiation of the most significant groups of methanogens regarding their shape and characteristic autofluorescent factor 420. Moreover, the fluorescence signal mirrors cell vitality. In this study, four different BGPs were investigated. The results indicated stable process performance in the mesophilic BGPs and in the thermophilic reactor. Bacterial subcommunity characterization revealed significant differences between the four BGPs. Most remarkably, the genera Defluviitoga and Halocella dominated the thermophilic bacterial subcommunity, whereas members of another taxon, Syntrophaceticus, were found to be abundant in the mesophilic BGP. The domain Archaea was dominated by the genus Methanoculleus in all four BGPs, followed by Methanosaeta in BGP1 and BGP3. In contrast, Methanothermobacter members were highly abundant in the thermophilic BGP4. Furthermore, a high consistency between the sequencing approach and the QMF method was shown, especially for the thermophilic BGP. The differences elucidated that using this biphasic approach for mesophilic BGPs provided novel insights regarding disaggregated single cells of Methanosarcina and Methanosaeta species. Both dominated the archaeal subcommunity and replaced coccoid Methanoculleus members belonging to the same group of Methanomicrobiales that have been frequently observed in similar BGPs. This work demonstrates that combining QMF and 16S rRNA gene amplicon sequencing is a complementary strategy to describe archaeal community structures within biogas processes.

• Journal of Microbiology and Biotechnology
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• v.16 no.5
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• pp.748-756
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• 2006

We propose a genome-scale clustering approach to identify whole genome relationships using the functional groups given by the Kyoto Encyclopedia of Genes and Genomes Orthology (KO) database. The metabolic capabilities of each organism were defined by the number of genes in each functional category. The archaeal, bacterial, and eukaryotic genomes were compared by simultaneously applying a two-step clustering method, comprised of a self-organizing tree algorithm followed by unsupervised hierarchical clustering. The clustering results were consistent with various phenotypic characteristics of the organisms analyzed and, additionally, showed a different aspect of the relationship between genomes that have previously been established through rRNA-based comparisons. The proposed approach to collect and cluster the metabolic functional capabilities of organisms should make it a useful tool in predicting relationships among organisms.

• Journal of Life Science
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• v.30 no.9
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• pp.813-818
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• 2020

The archaeal clusters of orthologous genes (arCOG) algorithm, which identifies common genes among archaebacterial genomes, was used to identify conservative genes among 168 archaebacterial strains. The numbers of conserved orthologs were 14, 10, 9, and 8 arCOGs in 168, 167, 166, and 165 strains, respectively. Among 41 conserved arCOGs, 13 were related to function J (translation, ribosomal structure, and biogenesis), and 10 were related to function L (replication, recombination, and repair). Among the 14 conserved arCOGs in all 168 strains, 6 arCOGs of tRNA synthetase comprised the highest proportion. Of the remaining 8 arCOGs, 2 are involved in reactions with ribosomes, 2 for tRNA synthesis, 2 for DNA replication, and 2 for transcription. These results showed the importance of protein expression in archaea. For the classes or orders having 3 or more members, genomic analysis was performed by averaging the distance values of the conservative arCOGs. Classes Archaeoglobi and Thermoplasmata of the phylum Euryarchaeota showed the lowest and the highest average of distance value, respectively. This study can provides data necessary for basic scientific research and the development of antibacterial agents and tumor control.

• Genomics & Informatics
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• v.8 no.4
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• pp.170-176
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• 2010

The Ribonucleotide reductases (RNR) are essential enzymes that catalyze the conversion of nucleotides to deoxynucleotides in DNA replication and repair in all living organisms. The RNRs operate by a free radical mechanism but differ in the composition of subunit, cofactor required and regulation by allostery. Based on these differences the RNRs are classified into three classesclass I, class II and class III which depend on oxygen, adenosylcobalamin and S-adenosylmethionine with an iron sulfur cluster respectively for radical generation. In this article thirty seven sequences belonging to each of the three classes of RNR were analyzed by using various tools of bioinformatics. Phylogenetic analysis, dot-plot comparisons and motif analysis was done to identify a number of differences in the three classes of RNRs. In this research article, we have attempted to decipher evolutionary relationship between the three classes of RNR by using bioinformatics approach.

• Journal of Photoscience
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• v.9 no.3
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• pp.45-48
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• 2002

Microbial rhodopsins, photoactive 7-transmembrane helix proteins that use retinal as their chromophore, were observed initially in the Archaea and appeared to be restricted to extreme halophilic environments. Our understanding of the abundance and diversity of this family has been radically transformed by findings over the past three years. Genome sequencing of cultivated microbes as well as environmental genomics have unexpectedly revealed archaeal rhodopsin homologs in the other two domains of life as well, namely Bacteria and Eucarya. Organisms containing these homologs inhabit such diverse environments as salt flats, soil, freshwater, and surface and deep ocean waters, and they comprise a broad phylogenetic range of microbial life, including haloarchaea, proteobacteria, cyanobacteria, fungi, and algae. Analysis of the new microbial rhodopsins and their expression and structural and functional characterization reveal that they fulfill both ion transport and sensory functions in various organisms, and use a variety of signaling mechanisms. We have obtained the first crystallographic structure for a photosensory member of this family, the phototaxis receptor sensory rhodopsin II (SRII, also known as phoborhodopsin) that mediates blue-light avoidance by the haloarchaeon Natronobacterium pharaonis. The structure obtained from x-ray diffraction of 3D crystals prepared in a cubic lipid phase reveals key features responsible for its spectral tuning and its sensory function. The mechanism of SRII signaling fits a unified model for transport and signaling in this widespread family of phototransducers.

• BMB Reports
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• v.38 no.6
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• pp.676-682
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• 2005

To date, no 8-oxoguanine-specific endonuclease-coding gene has been identified in Thermotoga maritima of the order Thermotogales, although its entire genome has been deciphered. However, the hypothetical protein Tm1821 from T. maritima, has a helix-hairpin-helix motif that is considered to be important for DNA binding and catalytic activity. Here, Tm1821 was overexpressed in Escherichia coli and purified using Ni-NTA affinity chromatography, protease digestion, and gel filtration. Tm1821 protein was found to efficiently cleave an oligonucleotide duplex containing 8-oxoguanine, but Tm1821 had little effect on other substrates containing modified bases. Moreover, Tm1821 strongly preferred DNA duplexes containing an 8-oxoguanine:C pair among oligonucleotide duplexes containing 8-oxoguanine paired with four different bases (A, C, G, or T). Furthermore, Tm1821 showed AP lyase activity and Schiff base formation with 8-oxoguanine in the presence of $NaBH_4$, which suggests that it is a bifunctional DNA glycosylase. Tm1821 protein shares unique conserved amino acids and substrate specificity with an 8-oxoguanine DNA glycosylase from the hyperthermophilic archaeon. Thus, the DNA recognition and catalytic mechanisms of Tm1821 protein are likely to be similar to archaeal repair protein, although T. maritima is an eubacterium.

• Journal of Microbiology and Biotechnology
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• v.15 no.6
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• pp.1359-1367
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• 2005

The gene encoding Pyrobaculum arsenaticum DNA polymerase (Par DNA polymerase) was cloned and sequenced. The gene consists of 2,361 bp coding for a protein with 786 amino acid residues. The deduced amino acid sequence of Par DNA polymerase showed a high similarity to archaeal family B-type DNA polymerases (Group I), and contained all of the motifs conserved in the family B-type DNA polymerases for $3'{\rightarrow}5'$ exonuclease and polymerase activities. The Par DNA polymerase gene was expressed under the control of the T7lac promoter on the expression vector pET-22b(+) in Escherichia coli BL21-CodonPlus(DE3)-RP. The expressed enzyme was purified by heat treatment, and Cibacron blue 3GA and $Hirap^{TM}$ Heparin HP column chromatographies. The optimum pH of the purified enzyme was 7.5. The enzyme activity was activated by divalent cations, and was inhibited by EDTA and monovalent cations. The half-life of the enzyme at $95^{\circ}C$ was 6 h. Par DNA polymerase possessed associated $3'{\rightarrow}5'$ proofreading exonuclease activity, which is consistent with its deduced amino acid sequence. PCR experiment with Par DNA polymerase showed an amplified product, indicating that this enzyme might be useful in DNA amplification and PCR-based applications.

• Microbiology and Biotechnology Letters
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• v.44 no.2
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• pp.171-179
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• 2016

Recent succession of soil microorganisms and vegetation has occurred in the glacier foreland, because of glacier thawing. In this study, whole microbial communities, including bacteria, archaea, and eukaryotes, from the glacier foreland of Midtre Lovénbreen in Svalbard were analyzed by metagenome sequencing, using the Ion Torrent Personal Genome Machine (PGM) platform. Soil samples were collected from two research sites (ML4 and ML7), with different exposure times, from the ice. A total of 2,798,108 and 1,691,859 reads were utilized for microbial community analysis based on the metagenomic sequences of ML4 and ML7, respectively. The relative abundance of microbial communities at the domain level showed a high proportion of bacteria (about 86−87%), whereas archaeal and eukaryotic communities were poorly represented by less than 1%. The remaining 12% of the sequences were found to be unclassified. Predominant bacterial groups included Proteobacteria (40.3% from ML4 and 43.3% from ML7) and Actinobacteria (22.9% and 24.9%). Major groups of Archaea included Euryarchaeota (84.4% and 81.1%), followed by Crenarchaeota (10.6% and 13.1%). In the case of eukaryotes, both ML4 and ML7 samples showed Ascomycota (33.8% and 45.0%) as the major group. These findings suggest that metagenome analysis using the Ion Torrent PGM platform could be suitably applied to analyze whole microbial community structures, providing a basis for assessing the relative importance of predominant groups of bacterial, archaeal, and eukaryotic microbial communities in the Arctic glacier foreland of Midtre Lovénbreen, with high resolution.