• Genomics & Informatics
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• v.21 no.1
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• pp.5.1-5.13
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• 2023

Neisseria gonorrhoeae is a Gram-negative aerobic diplococcus bacterium that primarily causes sexually transmitted infections through direct human sexual contact. It is a major public health threat due to its impact on reproductive health, the widespread presence of antimicrobial resistance, and the lack of a vaccine. In this study, we used a bioinformatics approach and performed subtractive genomic methods to identify potential drug targets against the core proteome of N. gonorrhoeae (12 strains). In total, 12,300 protein sequences were retrieved, and paralogous proteins were removed using CD-HIT. The remaining sequences were analyzed for non-homology against the human proteome and gut microbiota, and screened for broad-spectrum analysis, druggability, and anti-target analysis. The proteins were also characterized for unique interactions between the host and pathogen through metabolic pathway analysis. Based on the subtractive genomic approach and subcellular localization, we identified one cytoplasmic protein, 2Fe-2S iron-sulfur cluster binding domain-containing protein (NGFG RS03485), as a potential drug target. This protein could be further exploited for drug development to create new medications and therapeutic agents for the treatment of N. gonorrhoeae infections.

• Microbiology and Biotechnology Letters
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• v.29 no.1
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• pp.1-11
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• 2001

Lactic acid bacteria (LAB) are widely used for various food fermentation. With the recent advances in modern biotechnology, a variety of bio-products with the high economic values have been produced using microorganisms. For molecular cloning and expression studies on the gene of interest, E. coli has been widely used mainly because vector systems are fully developed. Most plasmid vectors currently used for E, coli carry antibiotic-resistant markers. As it is generally believed that the antibiotic resistance markers are potentially transferred to other bacteria, application of the plasmid vectors carrying antibiotic resistance genes as selection markers should be avoided, especially for human consump-tion. By contrast, as LAB have some desirable traits such that the they are GRAS(generally recognized as safe), able to secrete gene products out of cell, and their low protease activities, they are regarded as an ideal organism for the genetic manipulation, including cloning and expression of homologous and heterologous genes. However, the vec-tor systems established for LAB are stil insufficient to over-produce gene products, stably, limiting the use of these organisms for industrial applications. For a past decade, the two popular plasmid vectors, pAM$\beta$1 of Streptococcus faecalis and pGK12 theB. subtilis-E. coli shuttle vector derived from pWV01 of Lactococcus lactis ssp. cremoris wg 2, were most widely used to construct efficient chimeric vectors to be stably maintained in many industrial strains of LAB. Currently, non-antibiotic markers such as nisin resistance($Nis^{r}$ ) are explored for selecting recombi-nant clone. In addition, a gene encoding S-layer protein, slp/A, on bacterial cell wall was successfully recombined with the proper LAB vectors LAB vectors for excretion of the heterologous gene product from LAB Many food-grade host vec-tor systems were successfully developed, which allowed stable integration of multiple plasmid copies in the vec-mosome of LAB. More recently, an integration vector system based on the site-specific integration apparatus of temperate lactococcal bacteriophage, containing the integrase gene(int) and phage attachment site(attP), was pub-lished. In conclusion, when various vector system, which are maintain stably and expressed strongly in LAB, are developed, lost of such food products as enzymes, pharmaceuticals, bioactive food ingredients for human consump-tion would be produced at a full scale in LAB.

• BMB Reports
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• v.38 no.5
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• pp.507-516
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• 2005

Antimicrobial peptides (AMPs) have been isolated and characterized from tissues and organisms representing virtually every kingdom and phylum. Their amino acid composition, amphipathicity, cationic charge, and size allow them to attach to and insert into membrane bilayers to form pores by 'barrel-stave', 'carpet' or 'toroidal-pore' mechanisms. Although these models are helpful for defining mechanisms of AMP activity, their relevance to resolving how peptides damage and kill microorganisms still needs to be clarified. Moreover, many AMPs employ sophisticated and dynamic mechanisms of action to carry out their likely roles in antimicrobial host defense. Recently, it has been speculated that transmembrane pore formation is not the only mechanism of microbial killing by AMPs. In fact, several observations suggest that translocated AMPs can alter cytoplasmic membrane septum formation, reduce cell-wall, nucleic acid, and protein synthesis, and inhibit enzymatic activity. In this review, we present the structures of several AMPs as well as models of how AMPs induce pore formation. AMPs have received special attention as a possible alternative way to combat antibiotic-resistant bacterial strains. It may be possible to design synthetic AMPs with enhanced activity for microbial cells, especially those with antibiotic resistance, as well as synergistic effects with conventional antibiotic agents that lack cytotoxic or hemolytic activity.

• Journal of Microbiology and Biotechnology
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• v.34 no.6
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• pp.1276-1286
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• 2024

The environment has been identified as an origin, reservoir, and transmission route of antibiotic resistance genes (ARGs). Among diverse environments, freshwater environments have been recognized as pivotal in the transmission of ARGs between opportunistic pathogens and autochthonous bacteria such as Aeromonas spp. In this study, five environmental strains of Aeromonas spp. exhibiting multidrug resistance (MDR) were selected for whole-genome sequencing to ascertain their taxonomic assignment at the species-level and to delineate their ARG repertoires. Analyses of their genomes revealed the presence of one protein almost identical to AhQnr (A. hydrophila Qnr protein) and four novel proteins similar to AhQnr. To scrutinize the classification and taxonomic distribution of these proteins, all Aeromonas genomes deposited in the NCBI RefSeq genome database (1,222 genomes) were investigated. This revealed that these Aeromonas Qnr (AQnr) proteins are conserved intrinsic resistance determinants of the genus, exhibiting species-specific diversity. Additionally, structure prediction and analysis of contribution to quinolone resistance by AQnr proteins of the isolates, confirmed their functionality as quinolone resistance determinants. Given the origin of mobile qnr genes from aquatic bacteria and the crucial role of Aeromonas spp. in ARG dissemination in aquatic environments, a thorough understanding and strict surveillance of AQnr families prior to the clinical emergence are imperative. In this study, using comparative genome analyses and functional characterization of AQnr proteins in the genus Aeromonas, novel Aeromonas ARGs requiring surveillance has suggested.

• International Journal of Oral Biology
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• v.38 no.2
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• pp.61-65
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• 2013

Erythromycin is a macrolide antibiotic and inhibits bacterial protein synthesis by stimulating the dissociation of the peptidyl-tRNA molecule from the ribosomes during elongation. The use of macrolides has increased dramatically over the last few years and has led to an increase in bacterial resistance to these antibiotics. Bacterial resistance to erythromycin is generally conferred by the ribosome methylation and/or transport (efflux) protein genes. Among the identified erythromycin-resistant genes, erm(B) (erythromycin methylation) and mef(A) (macrolide efflux) are generally detectable in erythromycin-resistant streptococcal species. The distribution of these genes in oral streptococcal isolates has been reported in studies from other countries but has not been previously examined in a Korean study. We here examined by PCR the presence of erm(B) and mef(A) in oral streptococci isolated from Korean dental plaques. Among the 57 erythromycin-resistant strains tested, 64.9% harbored erm(B) whereas 40.4% were positive for mef(A). Eleven isolates had both the erm(B) and mef(A) genes. Twenty six isolates had only erm(B) and 12 isolates had only mef(A). Eight of the 57 strains examined were negative for both genes.

• Journal of Microbiology and Biotechnology
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• v.29 no.3
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• pp.500-505
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• 2019

Staphylococcus aureus is a round-shaped, gram-positive bacterium that can cause numerous infectious diseases ranging from mild infections such as skin infections and food poisoning to life-threatening infections such as sepsis, endocarditis and toxic shock syndrome. Various antibiotic-resistant strains of S. aureus have frequently emerged, threatening human lives significantly. Despite much research on the genetics of S. aureus, many of its genes remain unknown functionally and structurally. To counteract its toxins and to prevent the antibiotic resistance of S. aureus, our understanding of S. aureus should be increased at the proteomic scale. SAV0927 was first sequenced in an antibiotic resistant S. aureus strain. The gene is a conserved hypothetical protein, and its homologues appear to be restricted to Firmicutes. In this study, we determined the crystal structure of SAV0927 at $2.5{\AA}$ resolution. The protein was primarily dimeric both in solution and in the crystals. The asymmetric unit contained five dimers that are stacked linearly with ${\sim}80^{\circ}$ rotation by each dimer, and these interactions further continued in the crystal packing, resulting in a long linear polymer. The crystal structures, together with the network analysis, provide functional implications for the SAV0927-mediated protein network.

• Korean Journal of Microbiology
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• v.24 no.3
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• pp.263-269
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• 1986

Membrane proteins of facultatively anaerobic Escherichia coli K12 which was logarithmically grown in aerobiosis and anaerobiosis were compared on 5 to 10% liner gradient gel electrophoresis (Na Dod $SO_4 -PAGE$). Membrane proteins were formed as different patterns between aerobiosis and anaerobiosis. Among them, 91Kdal protein (pbp1a) was not synthesized in aerobiosis and 60Kdal protein (fts cluster), in anaerobiosis. Thereby cells cultured aerobically were differenciated as diversiform cell shape, comparing cells cultured anaerobically and the latter were resistant to penicillin G. Thus it is believed that in facultative anaerobes atmospheric oxygen regulated the synthesis of membrane proteins and even the expression of equivalent genes, and moreover alleviated the resistance to an antibiotic penicillin.

• Proceedings of the PSK Conference
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• 2001.04a
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• pp.180.3-181
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• 2001

• Journal of Microbiology and Biotechnology
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• v.34 no.8
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• pp.1718-1726
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• 2024

Development of novel antibacterial agents is imperative due to the increasing threat of antibiotic-resistant pathogens. This study aimed to develop the enhanced antibacterial activity and in-vivo efficacy of a novel truncated endolysin, CHAP^SAP26-161, derived from the endolysin Lys^SAP26, against multidrug-resistant bacteria. CHAP^SAP26-161 exhibited higher protein purification efficiency in E. coli and antibacterial activity than Lys^SAP26. Moreover, CHAP^SAP26-161 showed the higher lytic activity against A. baumannii with minimal bactericidal concentrations (MBCs) of 5-10 ㎍/ml, followed by Staphylococcus aureus with MBCs of 10-25 ㎍/ml. Interestingly, CHAP^SAP26-161 could lyse anaerobic bacteria, such as Clostridioides difficile, with MBCs of 25-50 ㎍/ml. At pH 4-8 and temperatures of 4℃-45℃, CHAP^SAP26-161 maintained antibacterial activity without remarkable difference. The lytic activity of CHAP^SAP26-161 was increased with Zn²⁺. In vivo tests demonstrated the therapeutic effects of CHAP^SAP26-161 in murine systemic A. baumannii infection model. In conclusion, CHAP^SAP26-161, a truncated endolysin that retains only the CHAP domain from Lys^SAP26, demonstrated enhanced protein purification efficiency and antibacterial activity compared to Lys^SAP26. It further displayed broad-spectrum antibacterial effects against S. aureus, A. baumannii, and C. difficile. Our in vitro and in-vivo results of CHAP^SAP26-161 highlights its promise as an innovative therapeutic option against those bacteria with multiple antibiotic resistance.

• Molecules and Cells
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• v.43 no.4
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• pp.350-359
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• 2020

Pathogenic aminoacyl-tRNA synthetases (ARSs) are attractive targets for anti-infective agents because their catalytic active sites are different from those of human ARSs. Mupirocin is a topical antibiotic that specifically inhibits bacterial isoleucyl-tRNA synthetase (IleRS), resulting in a block to protein synthesis. Previous studies on Thermus thermophilus IleRS indicated that mupirocin-resistance of eukaryotic IleRS is primarily due to differences in two amino acids, His581 and Leu583, in the active site. However, without a eukaryotic IleRS structure, the structural basis for mupirocin-resistance of eukaryotic IleRS remains elusive. Herein, we determined the crystal structure of Candida albicans IleRS complexed with Ile-AMP at 2.9 A resolution. The largest difference between eukaryotic and prokaryotic IleRS enzymes is closure of the active site pocket by Phe55 in the HIGH loop; Arg410 in the CP core loop; and the second Lys in the KMSKR loop. The Ile-AMP product is lodged in a closed active site, which may restrict its release and thereby enhance catalytic efficiency. The compact active site also prevents the optimal positioning of the 9-hydroxynonanoic acid of mupirocin and plays a critical role in resistance of eukaryotic IleRS to anti-infective agents.