Structural and biochemical analyses of the metallo-β-lactamase fold protein YhfI from Bacillus subtilis

Namgung, Byeol; Song, Wan Seok; Yoon, Sung‐il

doi:10.1016/j.bbrc.2019.08.106

Cited by 5 publications

(4 citation statements)

References 29 publications

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“…8a, b and Supplementary Table 2 ). Similar to other structurally characterized class II phosphodiesterases such as B. subtilis YhfI, yeast Saccharomyces cerevisiae PDE1 or widely distributed RNase Z proteins 21 , 22 , Apyc1 is a homodimer with a highly conserved HxHxDH motif that coordinates two Zn 2+ ions that bind phosphate groups to position cyclic nucleotides for cleavage (Extended Data Fig. 8a–c ).…”

Section: Mechanisms Of Cyclic Nucleotide Cleavagementioning

confidence: 95%

Phage anti-CBASS and anti-Pycsar nucleases subvert bacterial immunity

Hobbs

Wein

et al. 2022

Nature

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The cyclic oligonucleotide-based antiphage signalling system (CBASS) and the pyrimidine cyclase system for antiphage resistance (Pycsar) are antiphage defence systems in diverse bacteria that use cyclic nucleotide signals to induce cell death and prevent viral propagation1,2. Phages use several strategies to defeat host CRISPR and restriction-modification systems3–10, but no mechanisms are known to evade CBASS and Pycsar immunity. Here we show that phages encode anti-CBASS (Acb) and anti-Pycsar (Apyc) proteins that counteract defence by specifically degrading cyclic nucleotide signals that activate host immunity. Using a biochemical screen of 57 phages in Escherichia coli and Bacillus subtilis, we discover Acb1 from phage T4 and Apyc1 from phage SBSphiJ as founding members of distinct families of immune evasion proteins. Crystal structures of Acb1 in complex with 3′3′-cyclic GMP–AMP define a mechanism of metal-independent hydrolysis 3′ of adenosine bases, enabling broad recognition and degradation of cyclic dinucleotide and trinucleotide CBASS signals. Structures of Apyc1 reveal a metal-dependent cyclic NMP phosphodiesterase that uses relaxed specificity to target Pycsar cyclic pyrimidine mononucleotide signals. We show that Acb1 and Apyc1 block downstream effector activation and protect from CBASS and Pycsar defence in vivo. Active Acb1 and Apyc1 enzymes are conserved in phylogenetically diverse phages, demonstrating that cleavage of host cyclic nucleotide signals is a key strategy of immune evasion in phage biology.

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Section: Mechanisms Of Cyclic Nucleotide Cleavagementioning

confidence: 95%

Phage anti-CBASS and anti-Pycsar nucleases subvert bacterial immunity

Hobbs

Wein

et al. 2022

Nature

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“…Literature mining and phylogenetic analyses revealed that Rv1339, YfhI from Bacillus subtilis ( 29 ), and CpdA from Corynebacterium glutamicum ( 30 ) are representatives of a previously unrecognized class of PDEs defined by the signature metal-binding motif [T/S]HXHXDH, where X tends to be a hydrophobic, small or very small residue. This motif is reminiscent of the bona fide class-II PDE motif HXHLDH(27), and indeed, the two families are evolutionarily related—belonging to the metallohydrolase/oxidoreductase superfamily (SSF56281).…”

Section: Resultsmentioning

confidence: 99%

“…B , ribbon representation of a homology model of Rv1339 established using Modeller 9.23 ( 115 ) with the crystal structure of the metallo-β-lactamase fold protein YhfI from Bacillus subtilis (PDB: 6KNS ) as the template. YhfI was found to be a homodimer in solution by size-exclusion chromatography ( 29 ). The structure exhibits the alpha-beta-sandwich configuration characteristic of the metallo-β-lactamase fold, which consists of two β sheets of seven β strands each ( purple ) and α helices ( green ) capping the β sheet.…”

Section: Resultsmentioning

confidence: 99%

Expression of a novel mycobacterial phosphodiesterase successfully lowers cAMP levels resulting in reduced tolerance to cell wall–targeting antimicrobials

Liu

Nunta

Cheyne

et al. 2022

Journal of Biological Chemistry

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“…Literature mining and phylogenetic analyses revealed that Rv1339, YfhI from Bacillus subtilis 25 and CpdA from Corynebacterium glutamicum 26 , are representatives of a previously unrecognized class of PDEs defined by the signature metal-binding motif [T/S]HXHXDH, where X tends to be a hydrophobic, small or very small residue. This motif is reminiscent of the bona fide class-II PDE motif HXHLDH 27 , and indeed, both families are evolutionarily related and belong to the metallo-hydrolase/oxidoreductase superfamily (SSF56281).…”

Section: Resultsmentioning

confidence: 99%

Modulation of the cAMP levels with a conserved actinobacteria phosphodiesterase enzyme reduces antimicrobial tolerance in mycobacteria

Nunta

Cheyne

Liu

et al. 2020

Preprint

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Antimicrobial tolerance (AMT) is the gateway to the development of antimicrobial resistance (AMR) and is therefore a major issue that needs to be addressed.The second messenger cyclic-AMP (cAMP), which is conserved across all taxa, is involved in propagating signals from environmental stimuli and converting these into a response. In bacteria, such as M. tuberculosis, P. aeruginosa, V. cholerae and B. pertussis, cAMP has been implicated in virulence, metabolic regulation and gene expression. However, cAMP signalling in mycobacteria is particularly complex due to the redundancy of adenylate cyclases, which are enzymes that catalyse the formation of cAMP from ATP, and the poor activity of the only known phosphodiesterase (PDE) enzyme, which degrades cAMP into 5’- AMP.Based on these two features, the modulation of this system with the aim of investigating cAMP signalling and its involvement in AMT in mycobacteria id difficult.To address this pressing need, we identified a new cAMP-degrading phosphodiesterase enzyme (Rv1339) and used it to significantly decrease the intrabacterial levels of cAMP in mycobacteria. This analysis revealed that this enzyme increased the antimicrobial susceptibility of M. smegmatis mc2155. Using a combination of metabolomics, RNA-sequencing, antimicrobial susceptibility assays and bioenergetics analysis, we were able to characterize the molecular mechanism underlying this increased susceptibility.This work represents an important milestone showing that the targeting of cAMP signalling is a promising new avenue for antimicrobial development and expands our understanding of cAMP signalling in mycobacteria.

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Structural and biochemical analyses of the metallo-β-lactamase fold protein YhfI from Bacillus subtilis

Cited by 5 publications

References 29 publications

Phage anti-CBASS and anti-Pycsar nucleases subvert bacterial immunity

Phage anti-CBASS and anti-Pycsar nucleases subvert bacterial immunity

Expression of a novel mycobacterial phosphodiesterase successfully lowers cAMP levels resulting in reduced tolerance to cell wall–targeting antimicrobials

Modulation of the cAMP levels with a conserved actinobacteria phosphodiesterase enzyme reduces antimicrobial tolerance in mycobacteria

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