Characterization of Lhr-Core DNA helicase and manganese-dependent DNA nuclease components of a bacterial gene cluster encoding nucleic acid repair enzymes Journal Article
| Authors: | Ejaz, A.; Shuman, S. |
| Article Title: | Characterization of Lhr-Core DNA helicase and manganese-dependent DNA nuclease components of a bacterial gene cluster encoding nucleic acid repair enzymes |
| Abstract: | Lhr is a large superfamily 2 helicase present in mycobacteria and a moderate range of other bacterial taxa. A shorter version of Lhr, here referred to as Lhr-Core, is distributed widely in bacteria, where it is often encoded in a gene cluster along with predicted binuclear metallo-phosphoesterase (MPE), ATP-dependent DNA ligase, and metallo-β-lactamase exonuclease enzymes. Here we characterized the Lhr-Core and MPE proteins from Pseudomonas putida. We report that P. putida Lhr-Core is an ssDNA-dependent ATPase/dATPase (Km, 0.37 mM ATP; kcat, 3.3 s–1), an ATP-dependent 3'-to-5' single-stranded DNA translocase, and an ATP-dependent 3'-to-5' helicase. Lhr-Core unwinds 3'-tailed duplexes in which the loading/tracking strand is DNA and the displaced strand is either DNA or RNA. We found that P. putida MPE is a manganese-dependent phosphodiesterase that releases p-nitrophenol from bis-p-nitrophenyl phosphate (kcat, 212 s–1) and p-nitrophenyl-5'-thymidylate (kcat, 34 s-1) but displays no detectable phosphomonoesterase activity against p-nitrophenyl phosphate. MPE is also a manganese-dependent DNA endonuclease that sequentially converts a closed-circle plasmid DNA to nicked circle and linear forms prior to degrading the linear DNA to produce progressively smaller fragments. The biochemical activities of MPE and a structure predicted in Phyre2 point to MPE as a new bacterial homolog of Mre11. Genetic linkage of a helicase and DNA nuclease with a ligase and a putative exonuclease (a predicted homolog of the SNM1/Apollo family of nucleases) suggests that these enzymes comprise or participate in a bacterial DNA repair pathway. © 2018 Ejaz and Shuman. |
| Keywords: | dna; nucleic acids; enzymes; pseudomonas putida; gene encoding; repair; biochemical activity; bacteria; manganese; phosphodiesterases; phosphomonoesterase; single-stranded dna; dna repair pathways; signal encoding; bis-p-nitrophenyl phosphates; p-nitrophenyl phosphate |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 293 |
| Issue: | 45 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2018-11-09 |
| Start Page: | 17491 |
| End Page: | 17504 |
| Language: | English |
| DOI: | 10.1074/jbc.RA118.005296 |
| PUBMED: | 30224353 |
| PROVIDER: | scopus |
| PMCID: | PMC6231139 |
| DOI/URL: | |
| Notes: | Article -- Export Date: 3 December 2018 -- Source: Scopus |
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