Growth limiting substrate affects antibiotic production and associated metabolic fluxes in Streptomyces clavuligerus

Clavulanic acid biosynthesis by Streptomyces clavuligerus was dependent on the identity of the growth rate limiting nutrient in chemostat bioreactor culture (D=0.05 h(-1)). In phosphate-limited media, a specific production rate of 3.65 mg(clav) g(biomass) h(-1) was observed while N-limited media supported 0.32 mg(clav) g(biomass) h(-1). No production was observed in C-limited media. Metabolic flux analysis suggested that changing the nutrient limitation affected the availability of the C-5 precursor. Flux through anaplerotic metabolism was consistent with this, reflecting the lower rate of utilisation of 2-oxo-glutarate from the tricarboxylic acid (TCA) cycle for glutamate and, ultimately, C-5 precursor production, when antibiotic was not produced. We propose that C-limitation restricts the capacity for anaplerotic metabolism, minimising the potential for extensive TCA-cycle derived biosynthesis (the first stage in production of the C-5 precursor). N-Limitation would restrict the availability of nitrogen for amino acid biosynthesis (the next stage). Under P-limitation neither of these restrictions would apply.