Leveraging propionate-induced growth inhibition in Corynebacterium glutamicum to evolve improved methylmalonyl-CoA-dependent polyketide production

<title>Abstract</title> <italic>Corynebacterium glutamicum</italic> is a promising host for production of valuable polyketides. Propionate addition, a strategy known to increase polyketide production by increasing intracellular methylmalonyl-CoA availability, causes growth inhibition in <italic>C. glutamicum</italic>. The mechanism of this inhibition was unclear prior to our work. Here we provide evidence that accumulation of propionyl- and methylmalonyl-CoA induces growth inhibition in <italic>C. glutamicum</italic>. We then show that growth inhibition can be relieved by introducing methylmalonyl-CoA-dependent polyketide synthases. With germicidin as an example, we used adaptive laboratory evolution (ALE) to leverage the fitness advantage of polyketide production in the presence of propionate to evolve improved germicidin production. Whole genome sequencing revealed mutations in germicidin synthase (Gcs), which improved germicidin titer, as well as mutations in citrate synthase, which effectively evolved the native glyoxylate pathway to a new methylcitrate pathway. Together, our results show that <italic>C. glutamicum</italic> is a capable host for polyketide production, and we can take advantage of propionate growth inhibition to drive titers higher by evolution.