A Bacteriophage DNA Mimic Protein Employs a Non-specific Strategy to Inhibit the Bacterial RNA Polymerase

Abstract

DNA mimicry by proteins is a strategy that employed by some proteins to occupy the binding sites of the DNA-binding proteins and deny further access to these sites by DNA. Such proteins have been found in bacteriophage, eukaryotic virus, prokaryotic, and eukaryotic cells to imitate non-coding functions of DNA. Here, we report another phage protein Gp44 from bacteriophage SPO1 of Bacillus subtilis, employing mimicry as part of unusual strategy to inhibit host RNA polymerase. Consisting of three simple domains, Gp44 contains a DNA binding motif, a flexible DNA mimic domain and a random-coiled domain. Gp44 is able to anchor to host genome and interact bacterial RNA polymerase via the β and β′ subunit, resulting in bacterial growth inhibition. Our findings represent a non-specific strategy that SPO1 phage uses to target different bacterial transcription machinery regardless of the structural variations of RNA polymerases. This feature may have potential applications like generation of genetic engineered phages with Gp44 gene incorporated used in phage therapy to target a range of bacterial hosts.

Phage SPO1 Protein Gp49 Is a Novel RNA Binding Protein That Is Involved in Host Iron Metabolism

“…However, gene products from adjacent operons shed light on its function. Among the neighboring genes whose functions have been annotated, gene product 44 is a non-specific RNAP binding protein, that is able to shut down RNA synthesis for both E. coli and B. subtilis protein [ 6 ]. In the neighboring operon 3, Gp46 is a host HU protein inhibitor whose overexpression causes the loss of nucleoid structure [ 7 ].…”

“…Recent advances in annotating these gene products revealed several interesting host hijacking strategies employed by the phage. For example, Gp44 is a non-specific transcription inhibitor which contains a DNA binding motif that binds to DNA and a DNA mimic domain that traps RNA polymerase (RNAP) [ 6 ]. Gp46 is a cross-species HU inhibitor that causes filamentous morphology and nucleoid deformation in B. subtilis [ 7 ].…”

DNA-binding and protein structure of nuclear factors likely acting in genetic information processing in the Paulinella chromatophore

“… 27 ) or the DNA-binding N terminus of GP44, a bacteriophage SPO1 protein described as an RNA polymerase inhibitor (pdb-code: 6L6V) ref. 28 ]. However, the spatial arrangement of the secondary structures (especially the orientation of the helices toward the three-stranded β-sheet) differ in these other proteins, as seen by the high rmsd values of 2.7 Å (over 35 Cα atoms) and 3.6 Å (over 44 Cα atoms), respectively, when overlaid with sCTP-23166 ( SI Appendix , Fig.…”

Bacteriophage protein PEIP is a potent Bacillus subtilis enolase inhibitor

“…Thus, a potential PEIP-mimic drug targeting this interface and requiring high nanomolar affinity (219 nM) would have advantage over SF2312-like drugs. Furthermore, the cross-species inhibitory effect of PEIP, which also can be found in two other SPO1 proteins, Gp44 and Gp46 (Wang et al, 2021;Zhang et al, 2022), could enable a PEIPmimic drug to act as a broader range anti-microbial.…”