Antibiotics are indispensable for the treatment of a wide array of bacterial infections by destroying or suppressing bacterial proliferation. However, the prolonged utilization, misuse, and overuse of antibiotics have induced numerous bacteria to undergo genetic mutations and evolve drug resistance. In 2019, an estimated 4.95 million deaths were associated with antimicrobial resistance. Of these, 1.27 million deaths were attributed to bacterial antimicrobial resistance^[1]. By the year 2050, antibiotic resistance is estimated to be responsible for 10 million deaths globally, overtaking the predicted mortality from cancer^[2]. Antibiotic resistance has emerged as a leading cause of death and an urgent public health concern. Bacterial infections, such as pneumonia, tuberculosis, gonorrhea, and salmonellosis, are becoming increasingly difficult to treat due to proliferating drug resistance, leading to higher treatment costs, increased mortality, and rendering existing medication ineffectual^[3]. This calls for the continuous improvement of novel therapeutic agents as well as strategies to minimize the emergence of antibiotic resistance.

 

 

Figure 1 All-age rate of deaths attributable to and associated with bacterial antimicrobial resistance in 2019^[1].

 

To address the escalating dilemma of antibiotic resistance, our team opted to focus on antimicrobial peptides (AMPs), a class of small, endogenously produced peptides that inhibit diverse microorganisms. Compared to conventional antibiotics, AMPs exhibit lower toxicity and are less prone to induce resistance^[4]. However, natural AMPs are susceptible to enzymatic degradation and pH fluctuations, rendering them labile. They are also more costly to produce than antibiotics, and potentially toxic for oral therapy^[5]. Among them, nisin is an AMP generated by Lactococcus lactis, which displays potent activity against other Gram-positive species by attacking the cell wall and provoking lysis. Notably, nisin is the only bacteriocin approved as a food preservative, and it is usually used in dairy and meat products^[6]. Another AMP, darobactin, is produced by the bacterium Photorhabdus khanii. Darobactin represents the Gram-negative antibiotic with a novel scaffold, which induces cell lysis by disrupting the bacterial outer membrane^[7].

Given that both nisin and darobactin are ribosomally synthesized and post-translationally modified peptides (RiPPs) that follow the same biosynthetic logic. This project aims to combine them to produce a chimeric fusion peptide with complementary anti-Gram-positive and anti-Gram-negative biological activities. In this project, we genetically combined nisin with darobactin, expressed this engineered peptide with E. coli, and finally verified its activity. It will facilitate cost-effective, large-scale production of potent antimicrobial agents. In the future, we will formulate injectable preparations and topical ointments incorporating these peptides to treat bacterial infections in animals and humans.

 

Figure 2 Recombinant plasmids prepared based on combinatorial biosynthetic strategies.

 

 

Reference

[1] Murray C.J., Ikuta K.S., Sharara F., et al. Global burden of bacterial antimicrobial resistance in 2019: A systematic analysis [J]. Lancet, 2022, 399(10325): 629-655.

[2] Grabowicz M., Weiss D. Editorial overview: Antibiotics special issue [J]. Current Opinion in Microbiology, 2022, 65: V-VII.

[3] Organization W.H. Antibiotic resistance [Z]. 2020.https://www.who.int/news-room/fact-sheets/detail/antibiotic-resistance

[4] Rima M., Rima M., Fajloun Z., et al. Antimicrobial peptides: A potent alternative to antibiotics [J]. Antibiotics-Basel, 2021, 10(9): 1095.

[5] Bueno J., Demirci F., Baser K.H.C. Chapter 16 - antimicrobial strategies in novel drug delivery systems: Applications in the treatment of skin and soft tissue infections [M]. Kon K., Rai M. The microbiology of skin, soft tissue, bone and joint infections. Academic Press. 2017: 271-286.

[6] Gharsallaoui A., Oulahal N., Joly C., et al. Nisin as a food preservative: Part 1: Physicochemical properties, antimicrobial activity, and main uses [J]. Critical Reviews in Food Science and Nutrition, 2016, 56(8): 1262-1274.

[7] Imai Y., Meyer K.J., Iinishi A., et al. A new antibiotic selectively kills gram-negative pathogens [J]. Nature, 2019, 576(7787): 459-464.