Structural and Bioinformatics Analyses Identify Deoxydinucleotide-specific Nucleases and Their Association with Genomic Islands in Gram-positive Bacteria

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Date 2025 Jan 8

PMID 39778863

Authors

Sofia Mortensen

Stanislava Kuncova

Justin D Lormand

Tanner M Myers

Soo-Kyoung Kim

Vincent T Lee

Wade C Winkler

Holger Sondermann

Affiliations

Soon will be listed here.

Abstract

Dinucleases of the DEDD superfamily, such as oligoribonuclease, Rexo2 and nanoRNase C, catalyze the essential final step of RNA degradation, the conversion of di- to mononucleotides. The active sites of these enzymes are optimized for substrates that are two nucleotides long, and do not discriminate between RNA and DNA. Here, we identified a novel DEDD subfamily, members of which function as dedicated deoxydinucleases (diDNases) that specifically hydrolyze single-stranded DNA dinucleotides in a sequence-independent manner. Crystal structures of enzyme-substrate complexes reveal that specificity for DNA stems from a combination of conserved structural elements that exclude diribonucleotides as substrates. Consistently, diDNases fail to complement the loss of enzymes that act on diribonucleotides, indicating that these two groups of enzymes support distinct cellular functions. The genes encoding diDNases are found predominantly in genomic islands of Actinomycetes and Clostridia, which, together with their association with phage-defense systems, suggest potential roles in bacterial immunity.

References

Germain E, Castro-Roa D, Zenkin N, Gerdes K . Molecular mechanism of bacterial persistence by HipA. Mol Cell. 2013; 52(2):248-54. DOI: 10.1016/j.molcel.2013.08.045. View

Payne L, Todeschini T, Wu Y, Perry B, Ronson C, Fineran P . Identification and classification of antiviral defence systems in bacteria and archaea with PADLOC reveals new system types. Nucleic Acids Res. 2021; 49(19):10868-10878. PMC: 8565338. DOI: 10.1093/nar/gkab883. View

Doron S, Melamed S, Ofir G, Leavitt A, Lopatina A, Keren M . Systematic discovery of antiphage defense systems in the microbial pangenome. Science. 2018; 359(6379). PMC: 6387622. DOI: 10.1126/science.aar4120. View

Zuo Y, Wang Y, Malhotra A . Crystal structure of Escherichia coli RNase D, an exoribonuclease involved in structured RNA processing. Structure. 2005; 13(7):973-84. DOI: 10.1016/j.str.2005.04.015. View

Burastero O, Niebling S, Defelipe L, Gunther C, Struve A, Garcia Alai M . eSPC: an online data-analysis platform for molecular biophysics. Acta Crystallogr D Struct Biol. 2021; 77(Pt 10):1241-1250. PMC: 8489228. DOI: 10.1107/S2059798321008998. View

McCoy A, Grosse-Kunstleve R, Adams P, Winn M, Storoni L, Read R . Phaser crystallographic software. J Appl Crystallogr. 2009; 40(Pt 4):658-674. PMC: 2483472. DOI: 10.1107/S0021889807021206. View

Schneider C, Rasband W, Eliceiri K . NIH Image to ImageJ: 25 years of image analysis. Nat Methods. 2012; 9(7):671-5. PMC: 5554542. DOI: 10.1038/nmeth.2089. View

Altschul S, Gish W, Miller W, Myers E, Lipman D . Basic local alignment search tool. J Mol Biol. 1990; 215(3):403-10. DOI: 10.1016/S0022-2836(05)80360-2. View

Studier F . Protein production by auto-induction in high density shaking cultures. Protein Expr Purif. 2005; 41(1):207-34. DOI: 10.1016/j.pep.2005.01.016. View

10.

Prostova M, Kanevskaya A, Panteleev V, Lisitskaya L, Perfilova Tugaeva K, Sluchanko N . DNA-targeting short Argonautes complex with effector proteins for collateral nuclease activity and bacterial population immunity. Nat Microbiol. 2024; 9(5):1368-1381. DOI: 10.1038/s41564-024-01654-5. View

11.

Patel P, Maxwell K . Prophages provide a rich source of antiphage defense systems. Curr Opin Microbiol. 2023; 73:102321. DOI: 10.1016/j.mib.2023.102321. View

12.

Moser M, Holley W, Chatterjee A, Mian I . The proofreading domain of Escherichia coli DNA polymerase I and other DNA and/or RNA exonuclease domains. Nucleic Acids Res. 1998; 25(24):5110-8. PMC: 147149. DOI: 10.1093/nar/25.24.5110. View

13.

Georjon H, Tesson F, Shomar H, Bernheim A . Genomic characterization of the antiviral arsenal of Actinobacteria. Microbiology (Reading). 2023; 169(8). PMC: 10482375. DOI: 10.1099/mic.0.001374. View

14.

Millman A, Melamed S, Leavitt A, Doron S, Bernheim A, Hor J . An expanded arsenal of immune systems that protect bacteria from phages. Cell Host Microbe. 2022; 30(11):1556-1569.e5. DOI: 10.1016/j.chom.2022.09.017. View

15.

Bondy-Denomy J, Qian J, Westra E, Buckling A, Guttman D, Davidson A . Prophages mediate defense against phage infection through diverse mechanisms. ISME J. 2016; 10(12):2854-2866. PMC: 5148200. DOI: 10.1038/ismej.2016.79. View

16.

Alexander C, Wanner R, Johnson C, Breitsprecher D, Winter G, Duhr S . Novel microscale approaches for easy, rapid determination of protein stability in academic and commercial settings. Biochim Biophys Acta. 2014; 1844(12):2241-50. PMC: 4332417. DOI: 10.1016/j.bbapap.2014.09.016. View

17.

Goncalves O, de Assis J, Santana M . Breaking the ICE: an easy workflow for identifying and analyzing integrative and conjugative elements in bacterial genomes. Funct Integr Genomics. 2022; 22(6):1139-1145. DOI: 10.1007/s10142-022-00903-2. View

18.

Gerdes K, Baerentsen R, Brodersen D . Phylogeny Reveals Novel HipA-Homologous Kinase Families and Toxin-Antitoxin Gene Organizations. mBio. 2021; 12(3):e0105821. PMC: 8262856. DOI: 10.1128/mBio.01058-21. View

19.

Rousset F, Depardieu F, Miele S, Dowding J, Laval A, Lieberman E . Phages and their satellites encode hotspots of antiviral systems. Cell Host Microbe. 2022; 30(5):740-753.e5. PMC: 9122126. DOI: 10.1016/j.chom.2022.02.018. View

20.

Gao L, Altae-Tran H, Bohning F, Makarova K, Segel M, Schmid-Burgk J . Diverse enzymatic activities mediate antiviral immunity in prokaryotes. Science. 2020; 369(6507):1077-1084. PMC: 7985843. DOI: 10.1126/science.aba0372. View