Mechanism by which T7 bacteriophage protein Gp1.2 inhibits
            <i>Escherichia coli</i>
            dGTPase

Klemm, Bradley P.; Singh, Devender; Smith, Cassandra E.; Hsu, Allen; Dillard, Lucas; Krahn, J.M.; London, Robert E.; Mueller, Geoffrey A.; Borgnia, Mario J.; Schaaper, Roel M.

doi:10.1073/pnas.2123092119

Cited by 3 publications

(1 citation statement)

References 55 publications

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“…In turn, detected Ocr protein prevents the degradation and modification of phage DNA by inhibiting restriction endonuclease [ 74 , 75 ]. dGTPase inhibitor interacts and inhibits host dGTPase/dgt [ 76 ]. Two HNH endonuclease coding regions were detected in the Serratia phage KKP_3708 genome, which plays a key role in phage DNA packaging [ 77 ].…”

Section: Resultsmentioning

confidence: 99%

Biological Characterization and Genomic Analysis of Three Novel Serratia- and Enterobacter-Specific Virulent Phages

Shymialevich,

Błażejak,

Średnicka

et al. 2024

IJMS

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Due to the high microbiological contamination of raw food materials and the increase in the incidence of multidrug-resistant bacteria, new methods of ensuring microbiological food safety are being sought. One solution may be to use bacteriophages (so-called phages) as natural bacterial enemies. Therefore, the aim of this study was the biological and genomic characterization of three newly isolated Serratia- and Enterobacter-specific virulent bacteriophages as potential candidates for food biocontrol. Serratia phage KKP_3708 (vB_Sli-IAFB_3708), Serratia phage KKP_3709 (vB_Sma-IAFB_3709), and Enterobacter phage KKP_3711 (vB_Ecl-IAFB_3711) were isolated from municipal sewage against Serratia liquefaciens strain KKP 3654, Serratia marcescens strain KKP 3687, and Enterobacter cloacae strain KKP 3684, respectively. The effect of phage addition at different multiplicity of infection (MOI) rates on the growth kinetics of the bacterial hosts was determined using a Bioscreen C Pro growth analyzer. The phages retained high activity in a wide temperature range (from −20 °C to 60 °C) and active acidity values (pH from 3 to 12). Based on transmission electron microscopy (TEM) imaging and whole-genome sequencing (WGS), the isolated bacteriophages belong to the tailed bacteriophages from the Caudoviricetes class. Genomic analysis revealed that the phages have linear double-stranded DNA of size 40,461 bp (Serratia phage KKP_3708), 67,890 bp (Serratia phage KKP_3709), and 113,711 bp (Enterobacter phage KKP_3711). No virulence, toxins, or antibiotic resistance genes were detected in the phage genomes. The lack of lysogenic markers indicates that all three bacteriophages may be potential candidates for food biocontrol.

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Section: Resultsmentioning

confidence: 99%

Biological Characterization and Genomic Analysis of Three Novel Serratia- and Enterobacter-Specific Virulent Phages

Shymialevich,

Błażejak,

Średnicka

et al. 2024

IJMS

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Mechanism by which T7 bacteriophage protein Gp1.2 inhibits Escherichia coli dGTPase

Klemm

Singh

Smith

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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Levels of the cellular dNTPs, the direct precursors for DNA synthesis, are important for DNA replication fidelity, cell cycle control, and resistance against viruses. Escherichia coli encodes a dGTPase (2′-deoxyguanosine-5′-triphosphate [dGTP] triphosphohydrolase [dGTPase]; dgt gene, Dgt) that establishes the normal dGTP level required for accurate DNA replication but also plays a role in protecting E. coli against bacteriophage T7 infection by limiting the dGTP required for viral DNA replication. T7 counteracts Dgt using an inhibitor, the gene 1.2 product (Gp1.2). This interaction is a useful model system for studying the ongoing evolutionary virus/host “arms race.” We determined the structure of Gp1.2 by NMR spectroscopy and solved high-resolution cryo-electron microscopy structures of the Dgt–Gp1.2 complex also including either dGTP substrate or GTP coinhibitor bound in the active site. These structures reveal the mechanism by which Gp1.2 inhibits Dgt and indicate that Gp1.2 preferentially binds the GTP-bound form of Dgt. Biochemical assays reveal that the two inhibitors use different modes of inhibition and bind to Dgt in combination to yield enhanced inhibition. We thus propose an in vivo inhibition model wherein the Dgt–Gp1.2 complex equilibrates with GTP to fully inactivate Dgt, limiting dGTP hydrolysis and preserving the dGTP pool for viral DNA replication.

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Functionally comparable but evolutionarily distinct nucleotide-targeting effectors help identify conserved paradigms across diverse immune systems

Nicastro,

Burroughs,

Iyer

et al. 2023

Nucleic Acids Research

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While nucleic acid-targeting effectors are known to be central to biological conflicts and anti-selfish element immunity, recent findings have revealed immune effectors that target their building blocks and the cellular energy currency—free nucleotides. Through comparative genomics and sequence-structure analysis, we identified several distinct effector domains, which we named Calcineurin-CE, HD-CE, and PRTase-CE. These domains, along with specific versions of the ParB and MazG domains, are widely present in diverse prokaryotic immune systems and are predicted to degrade nucleotides by targeting phosphate or glycosidic linkages. Our findings unveil multiple potential immune systems associated with at least 17 different functional themes featuring these effectors. Some of these systems sense modified DNA/nucleotides from phages or operate downstream of novel enzymes generating signaling nucleotides. We also uncovered a class of systems utilizing HSP90- and HSP70-related modules as analogs of STAND and GTPase domains that are coupled to these nucleotide-targeting- or proteolysis-induced complex-forming effectors. While widespread in bacteria, only a limited subset of nucleotide-targeting effectors was integrated into eukaryotic immune systems, suggesting barriers to interoperability across subcellular contexts. This work establishes nucleotide-degrading effectors as an emerging immune paradigm and traces their origins back to homologous domains in housekeeping systems.

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Mechanism by which T7 bacteriophage protein Gp1.2 inhibits Escherichia coli dGTPase

Cited by 3 publications

References 55 publications

Biological Characterization and Genomic Analysis of Three Novel Serratia- and Enterobacter-Specific Virulent Phages

Biological Characterization and Genomic Analysis of Three Novel Serratia- and Enterobacter-Specific Virulent Phages

Mechanism by which T7 bacteriophage protein Gp1.2 inhibits Escherichia coli dGTPase

Functionally comparable but evolutionarily distinct nucleotide-targeting effectors help identify conserved paradigms across diverse immune systems

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