Transient analysis and engineering of bacterial metabolic pathways using cell-penetrating peptide-peptide nucleic acid conjugates

This study establishes CPP-PNAs as efficient tools for rapid, and simple metabolic pathway investigation. The approach produces results comparable to conventional genetic knockouts while offering dose-dependent control and avoiding permanent genomic alterations. Our findings reveal unexpected metabolic complexity in Synechocystis sp. PCC 6803 D-lactate synthesis under light conditions and demonstrate the utility of CPP-PNA for uncovering compensatory pathway activation. This platform represents a valuable addition to bacterial genetic engineering, addressing some of the critical limitations faced by conventional approaches, while showing potential for further understanding the biochemistry of metabolite-producing bacteria.