Journal of Microbiology

Journal Articles

Identification and Characterization of HEPN‑MNT Type II TA System from Methanothermobacter thermautotrophicus ΔH: Wonho Choi , Anoth Maharjan , Hae Gang Im , Ji-Young Park , Jong-Tae Park , Jung-Ho Park; J. Microbiol. 2023;61(4):411-421. Published online April 18, 2023; DOI: https://doi.org/10.1007/s12275-023-00041-9

Abstract: Toxin-antitoxin (TA) systems are widespread in bacteria and archaea plasmids and genomes to regulate DNA replication, gene transcr!ption, or protein translation. Higher eukaryotic and prokaryotic nucleotide-binding (HEPN) and minimal nucleotidyltransferase (MNT) domains are prevalent in prokaryotic genomes and constitute TA pairs. However, three gene pairs (MTH304/305, 408/409, and 463/464) of Methanothermobacter thermautotropicus ΔH HEPN-MNT family have not been studied as TA systems. Among these candidates, our study characterizes the MTH463/MTH464 TA system. MTH463 expression inhibited Escherichia coli growth, whereas MTH464 did not and blocked MTH463 instead. Using site-directed MTH463 mutagenesis, we determined that amino acids R99G, H104A, and Y106A from the R[ɸX]4-6H motif are involved with MTH463 cell toxicity. Furthermore, we established that purified MTH463 could degrade MS2 phage RNA, whereas purified MTH464 neutralized MTH463 activity in vitro. Our results indicate that the endonuclease toxin MTH463 (encoding a HEPN domain) and its cognate antitoxin MTH464 (encoding the MNT domain) may act as a type II TA system in M. thermautotropicus ΔH. This study provides initial and essential information studying TA system functions, primarily archaea HEPN-MNT family.

Analysis of Immune Responses Against Nucleocapsid Protein of the Hantaan Virus Elicited by Virus Infection or DNA Vaccination: Gyu-Jin Woo , Eun-Young Chun , Keun Hee Kim , Wankee Kim; J. Microbiol. 2005;43(6):537-545.; DOI: https://doi.org/2292 [pii]

Abstract: Even though neutralizing antibodies against the Hantaan virus (HTNV) has been proven to be critical against viral infections, the cellular immune responses to HTNV are also assumed to be important for viral clearance. In this report, we have examined the cellular and humoral immune responses against the HTNV nucleocapsid protein (NP) elicited by virus infection or DNA vaccination. To examine the cellular immune response against HTNV NP, we used H-2Kb restricted T-cell epitopes of NP. The NP-specific CD8+ T cell response was analyzed using a 51Cr-release assay, intracellular cytokine staining assay, enzyme-linked immunospot assay and tetramer binding assay in C57BL/6 mice infected with HTNV. Using these methods, we found that HTNV infection elicited a strong NP-specific CD8+ T cell response at eight days after infection. We also found that several different methods to check the NP-specific CD8+ T cell response showed a very high correlation among analysis. In the case of DNA vaccination by plasmid encoding nucleocapsid gene, the NP-specific antibody response was elicited 2 ~ 4 weeks after immunization and maximized at 6~8 weeks. NP-specific CD8+ T cell response reached its peak 3 weeks after immunization. In a challenge test with the recombinant vaccinia virus expressing NP (rVV-HTNV-N), the rVV-HTNV-N titers in DNA vaccinated mice were decreased about 100-fold compared to the negative control mice.

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