Systematic classification of the His-Me finger superfamily

Jabłońska, Jagoda; Matelska, Dorota; Steczkiewicz, Kamil; Ginalski, Krzysztof

doi:10.1093/nar/gkx924

Cited by 34 publications

(45 citation statements)

References 119 publications

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“…Our findings here strongly suggest that the P. aeruginosa EndA, like Dns and the S. oneidensis EndA, is an extracellular enzyme. Similar to other EndA orthologs, the P. aeruginosa EndA belongs to the family of ␤␤␣-metal endonucleases, also known as His-Me finger endonucleases, that form a large and diverse protein superfamily (70). The classification is based on a conserved structural motif, a ␤-hairpin followed by an ␣-helix, with a single metal ion being crucial for function (71).…”

Section: Discussionmentioning

confidence: 99%

Pseudomonas aeruginosa Requires the DNA-Specific Endonuclease EndA To Degrade Extracellular Genomic DNA To Disperse from the Biofilm

Cherny

Sauer

2019

J Bacteriol

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The dispersion of biofilms is an active process resulting in the release of planktonic cells from the biofilm structure. While much is known about the process of dispersion cue perception and the subsequent modulation of the c-di-GMP pool, little is known about subsequent events resulting in the release of cells from the biofilm. Given that dispersion coincides with void formation and an overall erosion of the biofilm structure, we asked whether dispersion involves degradation of the biofilm matrix. Here, we focused on extracellular genomic DNA (eDNA) due to its almost universal presence in the matrix of biofilm-forming species. We identified two probable nucleases, endA and eddB, and eddA encoding a phosphatase that were significantly increased in transcript abundance in dispersed cells. However, only inactivation of endA but not eddA or eddB impaired dispersion by Pseudomonas aeruginosa biofilms in response to glutamate and nitric oxide (NO). Heterologously produced EndA was found to be secreted and active in degrading genomic DNA. While endA inactivation had little effect on biofilm formation and the presence of eDNA in biofilms, eDNA degradation upon induction of dispersion was impaired. In contrast, induction of endA expression coincided with eDNA degradation and resulted in biofilm dispersion. Thus, released cells demonstrated a hyperattaching phenotype but remained as resistant to tobramycin as biofilm cells from which they egress, indicating EndA-dispersed cells adopted some but not all of the phenotypes associated with dispersed cells. Our findings indicate for the first time a role of DNase EndA in dispersion and suggest weakening of the biofilm matrix is a requisite for biofilm dispersion. IMPORTANCE The finding that exposure to DNase I impairs biofilm formation or leads to the dispersal of early stage biofilms has led to the realization of extracellular genomic DNA (eDNA) as a structural component of the biofilm matrix. However, little is known about the contribution of intrinsic DNases to the weakening of the biofilm matrix and dispersion of established biofilms. Here, we demonstrate for the first time that nucleases are induced in dispersed Pseudomonas aeruginosa cells and are essential to the dispersion response and that degradation of matrix eDNA by endogenously produced/secreted EndA is required for P. aeruginosa biofilm dispersion. Our findings suggest that dispersing cells mediate their active release from the biofilm matrix via the induction of nucleases.

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Section: Discussionmentioning

confidence: 99%

Pseudomonas aeruginosa Requires the DNA-Specific Endonuclease EndA To Degrade Extracellular Genomic DNA To Disperse from the Biofilm

Cherny

Sauer

2019

J Bacteriol

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“…Recently, SRA-HNH REases have been added to the list of modification-dependent, remote site cleaving, NTP-independent DNA endonucleases ( 36 ). The SRA-HNH REases are characterized by the presence of an SRA ( S ET and R ING finger a ssociated) domain ( 37 ) at the N-terminus and an HNH (histidine-asparagine-histidine) domain ( 38 ) at the C-terminus. Two enzymes from the family, ScoA3IV from Streptomyces coelicolor A3 (Sco5333) and TbiR51I from Thermobispora bispora (Tbis1) have been studied experimentally.…”

Section: Introductionmentioning

confidence: 99%

Crystal structure of the modification-dependent SRA-HNH endonuclease TagI

Kisiala

Copelas

Czapinska

et al. 2018

Nucleic Acids Research

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TagI belongs to the recently characterized SRA-HNH family of modification-dependent restriction endonucleases (REases) that also includes ScoA3IV (Sco5333) and TbiR51I (Tbis1). Here, we present a crystal structure of dimeric TagI, which exhibits a DNA binding site formed jointly by the nuclease domains, and separate binding sites for modified DNA bases in the two protomers. The nuclease domains have characteristic features of HNH/ββα-Me REases, and catalyze nicks or double strand breaks, with preference for /RY and RYN/RY sites, respectively. The SRA domains have the canonical fold. Their pockets for the flipped bases are spacious enough to accommodate 5-methylcytosine (5mC) or 5-hydroxymethylcytosine (5hmC), but not glucosyl-5-hydroxymethylcytosine (g5hmC). Such preference is in agreement with the biochemical determination of the TagI modification dependence and the results of phage restriction assays. The ability of TagI to digest plasmids methylated by Dcm (C5mCWGG), M.Fnu4HI (G5mCNGC) or M.HpyCH4IV (A5mCGT) suggests that the SRA domains of the enzyme are tolerant to different sequence contexts of the modified base.

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“…HNH domains are endonuclease domains that are very widespread in prokaryotes, but rarely found in eukaryotes (Jablonska et al, 2017). The domains are named after moderately conserved residues that were present in the founding members.…”

Section: Introductionmentioning

confidence: 99%

Protein Domain Guided Screen for Sequence Specific and Phosphorothioate-Dependent Restriction Endonucleases

et al. 2020

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Modification dependent restriction endonucleases (MDREs) restrict modified DNA, typically with limited sequence specificity (∼2-4 bp). Here, we focus on MDREs that have an SRA and/or SBD (sulfur binding domain) fused to an HNH endonuclease domain, cleaving cytosine modified or phosphorothioated (PT) DNA. We independently characterized the SBD-SRA-HNH endonuclease ScoMcrA, which preferentially cleaves 5hmC modified DNA. We report five SBD-HNH endonucleases, all recognizing GpsAAC/GpsTTC sequence and cleaving outside with a single nucleotide 3 stagger: EcoWI (N7/N6), Ksp11411I (N5/N4), Bsp305I (N6/N4-5), Mae9806I [N(8-10)/N(8-9)], and Sau43800I [N(8-9)/N(7-8)]. EcoWI and Bsp305I are more specific for PT modified DNA in Mg 2+ buffer, and promiscuous with Mn 2+. Ksp11411I is more PT specific with Ni 2+. EcoWI and Ksp11411I cleave fully-and hemi-PT modified oligos, while Bsp305I cleaves only fully modified ones. EcoWI forms a dimer in solution and cleaves more efficiently in the presence of two modified sites. In addition, we demonstrate that EcoWI PT-dependent activity has biological function: EcoWI expressing cells restrict dnd + GpsAAC modified plasmid strongly, and GpsGCC DNA weakly. This work establishes a framework for biotechnology applications of PT-dependent restriction endonucleases (PTDRs).

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Systematic classification of the His-Me finger superfamily

Cited by 34 publications

References 119 publications

Pseudomonas aeruginosa Requires the DNA-Specific Endonuclease EndA To Degrade Extracellular Genomic DNA To Disperse from the Biofilm

Pseudomonas aeruginosa Requires the DNA-Specific Endonuclease EndA To Degrade Extracellular Genomic DNA To Disperse from the Biofilm

Crystal structure of the modification-dependent SRA-HNH endonuclease TagI

Protein Domain Guided Screen for Sequence Specific and Phosphorothioate-Dependent Restriction Endonucleases

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