{"title":"Functional characterization of the mys genes for porphyra-334 biosynthesis from the terrestrial cyanobacterium Nostoc commune by heterologous expression.","authors":"Wei Yang, Toshio Sakamoto","doi":"10.2323/jgam.2025.05.002","DOIUrl":"https://doi.org/10.2323/jgam.2025.05.002","url":null,"abstract":"<p><p>Mycosporine-like amino acids (MAAs) are low-molecular-weight UV-protective compounds, and porphyra-334 and shinorine are common MAAs. Porphyra-334 is synthesized via the conjugation of mycosporine-glycine with threonine, whereas substitution with serine yields shinorine. The terrestrial cyanobacterium Nostoc commune KU002 (NIES-2538) produces 7-O-(β-arabinopyranosyl)-porphyra-334, and the mysABCD gene cluster responsible for MAA biosynthesis has been isolated. The heterologous expression of the mysABC genes from N. commune KU002 in Escherichia coli cells led to mycosporine-glycine production regardless of the culture medium supplemented with serine, threonine, or xylose. When the mysABCD genes from N. commune KU002 were expressed in E. coli cells, porphyra-334 production occurred, and shinorine production was observed upon serine supplementation in the culture medium. Notably, threonine and xylose supplementation in the culture medium increased the amounts of porphyra-334 in both cellular extracts and culture medium extracts. When the mysD gene was replaced with that from the shinorine producer Actinosynnema mirum JCM 3225, shinorine was primarily synthesized instead of porphyra-334. Interestingly, the transformant expressing the chimeric KU002-mysABC-JCM3225-mysD produced a novel MAA derivative with an absorption maximum at 334 nm and a molecular mass of 346 when cultured in the medium supplemented with threonine and xylose. These results suggest that the substrate specificity of MysD, which catalyzes the conjugation of mycosporine-glycine and serine or threonine, alters the production of porphyra-334 or shinorine and that the supplements added to the culture medium affect the amount and composition of MAAs produced in the E. coli transformant.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-05-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144199345","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Secretory Expression of a Multifunctional Nuclease Nuc A in Bacillus licheniformis 2709.","authors":"Huimin Guo, Kefen Wang, Tongtong Zhang, Honglei Fang, Wei Hui, Huitu Zhang","doi":"10.2323/jgam.2025.05.001","DOIUrl":"https://doi.org/10.2323/jgam.2025.05.001","url":null,"abstract":"<p><p>Serratia nuclease Nuc A is a non-specific nucleotide hydrolase that has been widely used in large-scale protein purification or eliminating nucleic acid contamination from purified proteins. To enhance the enzyme production, the Serratia nuclease gene was synthesized and expressed in Bacillus licheniformis 2709, a robust strain capable of secreting native and heterologous proteins selectively or non-selectively. To further increase the secretory expression level of the enzyme, different strong promoters and signal peptides were fused with the mature Nuc A-encoding gene at various genetic loci. The highest expression level of Nuc A was observed under the control of regulatory elements P<sub>aprE</sub>, which occur naturally in B. licheniformis 2709 for the alkaline protease (AprE) expression. Through maximizing the number of copies of P<sub>aprE-nucA</sub> expression cassette at different integration sites, the yield of nuclease Nuc A reached approximately 31954 U/mL after 60 hours of cultivation in shake flasks. The specific activity of the recombinant nuclease reached 1.58×107 U/mg, which is about 9 times higher than that expressed in Escherichia coli strain. Additionally, the recombinant Nuc A exhibited high catalytic activities in the pH range of 7-10. Furthermore, it was resistant to 0.2% SDS, 1.0 mM PMSF, and 0.4% Triton X-100. After 8 M Urea treatment, residual activity is measured. The high expression levels and positive characteristics of recombinant Nuc A provide an effective solution for large-scale production and industrial application of the nuclease.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-05-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144174138","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"PCR bias in 16S rRNA genes caused by GC content leads to insufficient detection of some abundant species in amplicon sequencing analyses of thermophilic microbial communities.","authors":"Shigeru Kawai, Katsumi Matsuura, Shawn Erin McGlynn","doi":"10.2323/jgam.2025.04.003","DOIUrl":"https://doi.org/10.2323/jgam.2025.04.003","url":null,"abstract":"<p><p>Amplicon sequencing is a widely used method for surveying biological diversity. However, the technique is disturbed by PCR bias leading to errors in community composition analyses. In this study, microbial community composition was evaluated in twenty-eight locations of hot spring water with temperatures between 87-48°C at Nakabusa Hot Springs, Japan, using amplicon sequencing analysis with the V4 region of the 16S rRNA gene. In discrepancy with the greenish color and the absorption spectra of the microbial samples, the relative abundance of amplicon sequence variants (ASVs) in the major photosynthetic organisms, Chloroflexus spp., were scarce in any sample when using the annealing temperature of 50°C in amplicon PCR. Changing the annealing temperature to 68ºC significantly improved the detection efficiency of Chloroflexus ASVs, and the obtained numbers were consistent with the presence of the photosynthetic pigments. The abundance of many other microbial ASVs was also dependent on the annealing temperature. The log ratio in the abundance of major ASVs between two annealing temperatures was correlated with the GC content of the 16S rRNA gene, suggesting that even some other major ASVs in the community are seriously affected by PCR bias due to the GC content. Combined usage of results from two different annealing temperatures, rather than a result using a single annealing temperature, seems to be a better way to obtain community structure information with less PCR bias in thermophilic organisms of high 16S rRNA GC content.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-05-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144142698","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Determination of double bond configuration of 2-hydroxy-fatty acids and emendation of cellular fatty acid composition of Aureispira marina and Aureispira maritima.","authors":"Fuka Iriyama, Hirokazu Iida, Kazuyoshi Kawahara","doi":"10.2323/jgam.2024.09.001","DOIUrl":"10.2323/jgam.2024.09.001","url":null,"abstract":"<p><p>Aureispira marina is a marine bacterium with gliding motility isolated from the southern coastline of Thailand. It contained ceramide as a major cellular lipid composed of saturated or unsaturated branched chain 2-hydroxy-fatty acid and sphingosine. The structure of unsaturated 2-hydroxy-fatty acid was investigated in our previous study, but the geometric configuration of the double bond remained unclear. In the present study, 14-methyl-∆<sup>2</sup>-pentadecenol (∆<sup>2</sup>-iso-C<sub>16:1</sub>-ol) was prepared from D-2-hydroxy-15-methyl-∆<sup>3</sup>-hexadecenoic acid (D-2-OH-∆<sup>3</sup>-iso-C<sub>17:1</sub>) of the ceramide component, and analyzed by <sup>1</sup>H and <sup>13</sup>C NMR in comparison with ∆<sup>2</sup>-trans-hexadecenol (∆<sup>2</sup>-trans-n-C<sub>16:1</sub>-ol) derived from commercially available D-sphingosine. From the coupling constants of protons in the double bond and the chemical shift value of allylic carbon, the configuration of the double bond was determined as trans. Since the structure of 2-hydroxy-fatty acids was clarified, cellular fatty acids of A. marina and A. maritima, another species of the genus Aureispira, were reexamined, and the description on the cellular fatty acid composition of the genus Aureispira in the previous papers (Hosoya et al., 2006, Int. J. System. Evol. Microbiol., 56, 2931-2935; Hosoya et al., 2007, Int. J. System. Evol. Microbiol., 57, 1948-1951) lacking the description of 2-hydroxy-fatty acids was emended.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142348068","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"A DUF3844 domain-containing protein is required for vacuolar protein sorting in Schizosaccharomyces pombe.","authors":"Tomoaki Inagawa, Kazuma Ohkubo, Masahiro Watanabe, Tomotake Morita, Yujiro Higuchi, Hiromi Maekawa, Kaoru Takegawa","doi":"10.2323/jgam.2024.10.001","DOIUrl":"10.2323/jgam.2024.10.001","url":null,"abstract":"<p><p>Protein trafficking to vacuoles in plants and fungi, and to lysosomes in animals, is essential for the maintenance of cellular homeostasis. In Saccharomyces cerevisiae, the vacuolar protein sorting (VPS) pathway has been well studied by using vacuolar carboxypeptidase Y (CPY) as a model, and many VPS genes have been identified. By contrast, the vacuolar protein trafficking pathway in Schizosaccharomyces pombe remains poorly understood. In this study, we identified a novel VPS gene (SPBC1709.03) in S. pombe that is broadly conserved in fungi, but not in S. cerevisiae. Owing to its DUF3844 domain of unknown function, the gene was named vps3844. Disruption mutants of vps3844 had defects in both CPY sorting and incorporation of FM4-64 dye into the vacuolar membrane. Partial deletion analysis of the Vps3844 protein revealed that, within the DUF3844 domain, the region comprising amino acids 354 to 380 is important for protein trafficking to the vacuole. Our findings represent the first report of a VPS gene involved in vacuolar transport that is conserved in fungi, particularly S. pombe, but lacks representation in S. cerevisiae.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142545832","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Fauzi Akhbar Anugrah, I Nyoman Pugeg Aryantha, Rahmi Masita, Siti Zubaidah, Nur Izzati Mohd Noh
{"title":"Isolation of Bacterial Endophytes Associated with Cinchona ledgeriana Moens. and Their Potential in Plant-growth Promotion, Antifungal and Quinoline Alkaloids Production.","authors":"Fauzi Akhbar Anugrah, I Nyoman Pugeg Aryantha, Rahmi Masita, Siti Zubaidah, Nur Izzati Mohd Noh","doi":"10.2323/jgam.2024.09.002","DOIUrl":"10.2323/jgam.2024.09.002","url":null,"abstract":"<p><p>For centuries, quinoline alkaloids from the tree bark of Cinchona ledgeriana (C. ledgeriana) have been used in the treatment of malaria. However, unsustainable harvesting and poor growth conditions greatly limit its use as raw materials. Since plant endophytes are known to contribute to the physiology of the host and its metabolism for survival, this study showed the potential of endophytes isolated from C. ledgeriana roots in promoting the germination of Catharathus roseus (C. roseus) seedlings and the biosynthesis of quinoline alkaloid. In this present study, we found that the Enterobacteriaceae family comprised the majority of the bacterial community, with Klebsiella pneumoniae being the most abundant species at the C. ledgeriana roots. Characterization of culturable bacterial endophytes from the C. ledgeriana roots showed that all the isolates displayed plant growth-promoting factors and antifungal activities. Interestingly, chromatographic analyses led to the identification of the quinoline alkaloids producing Achromobacter xylosoxidans (A. xylosoxidans) A1. Moreover, the co-cultures of A. xylosoxidans A1, Cytobacillus solani (C. solani) A3, and Klebsiella aerogenes A6 increased the fresh and dry weight of the C. roseus seedlings. These results suggest that these bacterial endophytes may enhance quinine and quinidine production as well as the growth of the plant host.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142501746","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Microbial community analysis of sand filters used to treat mine water from a closed uranium mine.","authors":"Hiroshi Habe, Tomohiro Inaba, Tomo Aoyagi, Hidenobu Aizawa, Yuya Sato, Tomoyuki Hori, Keiko Yamaji, Yoshiyuki Ohara, Kenjin Fukuyama, Takuro Nishimura","doi":"10.2323/jgam.2024.08.001","DOIUrl":"10.2323/jgam.2024.08.001","url":null,"abstract":"<p><p>Rapid sand filters (RSFs) are employed in a drinking water treatment to remove undesirable elements such as suspended solids and dissolved metal ions. At a closed uranium (U) mine site, two sets of tandemly linked paired RSF systems (RSF1-RSF2 and RSF1-RSF3) were utilized to remove iron and manganese from mine water. In this study, a 16S rRNA-based amplicon sequencing survey was conducted to investigate the core microbes within the RSF system treating the mine water. In RSF1, two operational taxonomic units (OTUs) related to methanotrophic bacteria, Methylobacter tundripaludum (relative abundance: 18.1%) and Methylovulum psychrotolerans (11.5%), were the most and second most dominant species, respectively, alongside iron-oxidizing bacteria. The presence of these OTUs in RSF1 can be attributed to the microbial community in the inlet mine water, as the three most abundant OTUs in the mine water also dominated RSF1. Conversely, in both RSF2 and RSF3, Nevskia sp., previously isolated from the Ytterby mine manganese oxide producing ecosystem, became dominant, although known manganese-oxidizing bacterial OTUs were not detected. In contrast, a unique OTU related to Rhodanobacter sp. was the third most abundant (8.0%) in RSF1, possibly due to selective pressure from the radionuclide-contaminated environment during RSF operation, as this genus is known to be abundant at nuclear legacy waste sites. Understanding the key bacterial taxa in RSF system for mine water treatment could enhance the effectiveness of RSF processes in treating mine water from closed U mines.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142289014","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Ran Deng, Jing Li, Bo Yu Liu, Jie Du, JianGuo Lu, Qiang Li, QianRu Hou
{"title":"Isolation and identification of naphtalene-degrading bacteria and its application in a two-phase partitioning bioreactor.","authors":"Ran Deng, Jing Li, Bo Yu Liu, Jie Du, JianGuo Lu, Qiang Li, QianRu Hou","doi":"10.2323/jgam.2024.07.003","DOIUrl":"10.2323/jgam.2024.07.003","url":null,"abstract":"<p><p>Naphthalene is a persistent environmental pollutant for its potential teratogenic, carcinogenic and mutagenic effects. In this study, 10 strains of bacteria capable of degrading naphthalene were isolated from crude-oil contaminated soil. Among them, Pseudomonas plecoglossicida 2P exhibited prominent growth with 1000 mg/L naphthalene as the sole carbon source and degraded 94.15% of naphthalene in 36 h. Whole genome sequencing analysis showed that P. plecoglossicida 2P had a total of 22 genes related to naphthalene degradation, of which 8 genes were related to the salicylic acid pathway only, 5 genes were related to the phthalic acid pathway only, 8 genes were common in both the salicylic acid and phthalic acid pathways, and 1 gene was related to the gentisic acid pathway. P. plecoglossicida 2P was applied in a two-phase partition bioreactor (TPPB) to degrade naphthalene in wastewater. The optimal operating conditions of the reactor were obtained through response surface optimization: initial naphthalene concentration (C<sub>0</sub>) =1600 mg/L, bacterial liquid concentration (OD<sub>600</sub>) = 1.3, and polymer-to-wastewater mass ratio (PWR) = 2%. Under these conditions, the naphthalene degradation rate was 98.36% at 24 h. The degradation kinetics were fitted using the Haldane equation with a high coefficient of determination (R<sup>2</sup>=0.94). The present study laid foundations for naphthalene degradation mechanism of genus Pseudomonas and its potential application in TPPB.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141971289","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"24R005A and 24R005B: Novel radical scavengers of DPPH obtained from Streptomyces sp. cultured in a fish powder medium.","authors":"Toshikazu Komoda, Mayu Abe, Yoshitaka Koseki","doi":"10.2323/jgam.2024.07.002","DOIUrl":"10.2323/jgam.2024.07.002","url":null,"abstract":"<p><p>We have successfully isolated two novel compounds, 24R005A (1, C<sub>13</sub>H<sub>14</sub>O<sub>4</sub>) and 24R005B (2, C<sub>13</sub>H<sub>13</sub>ClO<sub>4</sub>), from Streptomyces sp. 24R005, using fish (anchovy) powder as a medium. In this study, we evaluated the use of fish (anchovy) powder as a fermentation material for producing bioactive compounds. Spectroscopic analyses revealed that the two compounds share a common skeletal structure. However, each compound contains unique branched side chains. Furthermore, compounds 1 and 2 exhibit moderate radical-scavenging activity for 1,1-diphenyl-2-picrylhydrazyl (DPPH), with ED<sub>50</sub> values of 200 and 130 μM, respectively.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860015","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Marine bacteria have multiple polyamide 4-degrading enzymes.","authors":"Yusuke Saito, Ibuki Jin, Miwa Yamada","doi":"10.2323/jgam.2024.05.001","DOIUrl":"10.2323/jgam.2024.05.001","url":null,"abstract":"<p><p>Polyamide 4 (PA4) is expected to solve the issue of marine plastic pollution due to its excellent mechanical properties and biodegradability. In this study, to reveal the mechanism of PA4 biodegradation in the marine environment, we isolated 5 strains of PA4-degrading bacteria belonging to Aliiglaciecola, Dasania, and Pseudophaeobacter from a marine environment. The isolated 5 strains are novel PA4-degrading bacteria that are phylogenetically distinct from those isolated in previous studies. In addition, we compared the PA4-degrading activities and structures of the PA4-degrading enzymes secreted by the 5 strains and PA4-degrading strains isolated in our previous study. The PA4-degrading activity in the supernatant of the cultivation solutions differed among the strains. Native-PAGE and zymography using a polyacrylamide gel containing a PA4 emulsion demonstrated that PA4-degrading enzymes are classified into no less than three types of structures. These results suggested that marine PA4-degrading bacteria have multiple PA4-degrading enzymes. Our findings will contribute to a better understanding of the microbial degradation of PA4 in the marine environment.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":0.8,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140911066","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}