A structurally unique aminoglycoside produced in Streptoalloteichus tenebrarius, Apramycin is used in veterinary medicine or the treatment of Salmonella, Escherichia coli and Pasteurella multocida infections. Although apramycin was discovered nearly 50 years ago, many biosynthetic steps of apramycin remain unknown. In this study, we identified a HemK family methyltransferase, AprI, to be the 7’-N-methyltransferase in apramycin biosynthetic pathway. Biochemical experiments showed that AprI converted demethyl-aprosamine to aprosamine. Through gene disruption of aprI, we identified a new aminoglycoside antibiotic demethyl-apramycin as the main product in aprI disruption strain. The demethyl-apramycin is an impurity in apramycin product. In addition to demethyl-apramycin, carbamyltobramycin is another major impurity. However, unlike demethyl-apramycin, tobramycin is biosynthesized by an independent biosynthetic pathway in S. tenebrarius. The titer and rate of apramycin were improved by overexpression of the aprI and disruption of the tobM2, which is a crucial gene for tobramycin biosynthesis. The titer of apramycin increased from 2227±320 mg/L to 2331±210 mg/L, while the titer of product impurity demethyl-apramycin decreased from 196±36 mg/L to 51±9 mg/L. Moreover, the carbamyltobramycin titer of the wild-type strain was 607±111 mg/L and that of the engineering strain was null. The rate of apramycin increased from 68% to 87%, and that of demethyl-apramycin decreased from 1.17% to 0.34%.

Background Multidrug-resistant staphylococcus aureus infected wounds can lead to nonhealing, systemic infections, and even death. Although advanced dressings are effective in protecting, disinfecting, and maintaining moist microenvironments, they often have limitations such as single functionality, inadequate drug release, poor biosafety, or high rates of drug resistance. Methods Here, a novel wound dressing comprising glycyrrhizic acid (GA) and tryptophan-sorbitol carbon quantum dots (WS-CQDs) was developed, which exhibit synergistic and long-lasting antibacterial and anti-inflammatory effects. We investigated the characterization, mechanical properties, synergistic antibacterial effects, sustained-release properties, and cytotoxicity of GA/WS-CQDs hydrogels in vitro. Additionally, we performed transcriptome sequence analysis to elucidate the antibacterial mechanism. Furthermore, we evaluated the biosafety, anti-inflammatory effects, and wound healing ability of GA/WS-CQDs dressings using an in vivo mouse model of methicillin-resistant staphylococcus aureus (MRSA)-infected wounds. Results The prepared GA/WS-CQDs hydrogels demonstrated superior anti-MRSA effects compared to common antibiotics in vitro. Furthermore, the sustained release of WS-CQDs from GA/WS-CQDs hydrogels lasted for up to 60 h, with a cumulative release of exceeding 90%. The sustained-released WS-CQDs exhibited excellent anti-MRSA effects, with low drug resistance attributed to DNA damage and inhibition of bacterial biofilm formation. Notably, in vivo experiments showed that GA/WS-CQDs dressings reduced the expression of inflammatory factors (TNF-α, IL-1β, and IL-6) and significantly promoted the healing of MRSA-infected wounds with almost no systemic toxicity. Importantly, the dressings did not require replacement during the treatment process. Conclusion These findings emphasize the high suitability of GA/WS-CQDs dressings for MRSA-infected wound healing and their potential for clinical translation.

BackgroundApramycin is a structurally unique aminoglycoside, used in veterinary medicine or the treatment of Salmonella, Escherichia coli and Pasteurella multocida infections in farm. Although discovered and used many years ago, many biosynthetic steps of apramycin are still obscure. ResultsIn this study, we identified a HemK family methyltransferase, aprI, involved in apramycin biosynthesis. The function of aprI was studied by using gene disruption and biochemical experiments, and a new aminoglycoside antibiotic demethyl-apramycin was purified from aprI disruption strain. Experiments proved that AprI converted demethyl-aprosamine to aprosamine in vitro. Based on this, the apramycin production strain was improved by overexpression the AprI to decrease the impurity production. ConclusionsWe have identified aprI is a 7’-N-methyltransferase gene in apramycin biosynthesis and confirmed the substrate of methyltransferase. Engineering of aprI resulted in a strain producing a new aminoglycoside demethyl-apramycin and apramycin mono-producing strain with less impurity production. Finally, the yield of demethyl-apramycin in apramycin mono-producing strain decreased from 196±36 mg/L to 51±9 mg/L, and the yield of apramycin increased from 2227±320 mg/L to 2331±210 mg/L.