Mining for Novel Bioactive Fungal Metabolites through Precursor-directed Biosynthesis

Abstract

Antibiotics have played a crucial role in modern medicine for more than 60 years by effectively treating infectious diseases and reducing the occurrence of infections in immunocompromised patients. Nevertheless, the excessive use of antibiotics has resulted in an increase in antibiotic-resistant bacteria, requiring the development of new antibiotics with unique mechanisms. One promising approach is exploring natural products such as fungal secondary metabolites. Precursor-directed biosynthesis (PDB) involves the use of specific precursor molecules to produce modified metabolites possessing unique activities, potentially useful in medicine, agriculture, and other fields. PDB uses the adaptive metabolic processes of microorganisms to incorporate unnatural precursors, creating novel derivatives with potentially improved efficacy and reduced toxicity. This approach requires understanding microbial growth, precursor behaviour, and biosynthetic pathways. Advances in synthetic biology and metabolic engineering have further expanded these possibilities. Aspergillus species, commonly found fungi, can exhibit both advantageous and detrimental effects. Certain species can cause infections and producing harmful mycotoxins, whereas Aspergillus terreus is known for its ability to thrive in adverse conditions and produce beneficial secondary metabolites, including lovastatin (a cholesterol-lowering drug) and antibacterial agents like terretonin. In this study, the incorporation of phenylalanine and its derivatives into the biosynthetic pathways of A. terreus was investigated to enhance the production of modified aromatic metabolites. The study examined six phenylalanine derivatives: phenylalanine, dihydroxyphenylalanine, β-phenylalanine, fluoro-phenylalanine, chloro-phenylalanine, and bromo-phenylalanine. Each derivative was isolated and purified through similar extraction protocols, ensuring consistency and comparability in the resulting fractions. The fractions were then subjected to Minimum Inhibitory Concentration (MIC) assays to evaluate their antimicrobial efficacy against Staphylococcus aureus and Escherichia coli. Further chemical analysis was conducted using solid-phase extraction (SPE) and high-performance liquid chromatography (HPLC) to separate and identify the compounds present. This study demonstrates the potential of A. terreus in generating bioactive secondary metabolites through PDB, offering a promising strategy for the development of novel antimicrobial agents. The findings highlight the importance of investigating fungal metabolites and metabolic engineering to enhance the biosynthetic capacity of microorganisms, in response to the demand of new antibiotics due to rising antimicrobial resistance.