Identification of Gip as a novel phage‐encoded gyrase inhibitor protein of Corynebacterium glutamicum

• Published in: Molecular microbiology 2021-11, Vol.116 (5), p.1268-1280
• Main Authors: Kever, Larissa, Hünnefeld, Max, Brehm, Jannis, Heermann, Ralf, Frunzke, Julia
• Format: Article
• Language: English
• Subjects: Anti-Bacterial Agents - metabolism; Antiinfectives and antibacterials; bacteriophages; Corynebacterium glutamicum; Corynebacterium glutamicum - virology; Deoxyribonucleic acid; DNA; DNA biosynthesis; DNA gyrase; DNA Replication; DNA topoisomerase; Drug delivery; Drug development; GIP protein; gyrase inhibitors; High-Throughput Screening Assays - methods; Phages; prophage induction; Prophages - genetics; Prophages - metabolism; Proteins; RecA protein; SOS response; Surface plasmon resonance; topoisomerase II inhibitors; Topoisomerase II Inhibitors - metabolism; Topology; Viral Proteins - genetics; Viral Proteins - metabolism
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1111/mmi.14813

By targeting key regulatory hubs of their host, bacteriophages represent a powerful source for the identification of novel antimicrobial proteins. Here, a screening of small cytoplasmic proteins encoded by the CGP3 prophage of Corynebacterium glutamicum resulted in the identification of the gyrase‐inhibiting protein Cg1978, termed Gip. Pull‐down assays and surface plasmon resonance revealed a direct interaction of Gip with the gyrase subunit A (GyrA). The inhibitory activity of Gip was shown to be specific to the DNA gyrase of its bacterial host C. glutamicum. Overproduction of Gip in C. glutamicum resulted in a severe growth defect as well as an induction of the SOS response. Furthermore, reporter assays revealed an RecA‐independent induction of the cryptic CGP3 prophage, most likely caused by topological alterations. Overexpression of gip was counteracted by an increased expression of gyrAB and a reduction of topA expression at the same time, reflecting the homeostatic control of DNA topology. We postulate that the prophage‐encoded Gip protein plays a role in modulating gyrase activity to enable efficient phage DNA replication. A detailed elucidation of the mechanism of action will provide novel directions for the design of drugs targeting DNA gyrase. Bacteriophages represent a rich source for the identification of novel antimicrobial proteins. Here, we describe the identification and first characterization of a small prophage‐encoded protein Gip as a novel gyrase inhibitor specifically targeting the DNA gyrase of its bacterial host Corynebacterium glutamicum.