The rise of antibiotic resistant pathogens motivates research into new treatments to prevent the resurgence of infectious and deadly diseases. (GccF) a diglycosylated bacteriocin secreted by Lactobacillus plantarum KW30, completely inhibits the growth of susceptible cells within five minutes at low (1-5) nM concentrations in rich media using what appears to be a novel bacteriostatic mechanism. Understanding this mechanism could provide a blueprint for the design of a new family of antibiotics. Requirements for the biosynthesis of this peptide are unknown as attempts to heterologously express or modify the seven genes gccA-F and gccH within the cluster have been unsuccessful, preventing experimental verification of their activities. Here we report the development of an easily modifiable 11.2 kbp plasmid based heterologous expression system producing active GccF in L. plantarum NC8 and L. Sakei 790 that has revolutionised research into its maturation pathway. Expression relies on the promotors found naturally within the cluster and produces the same concentration of peptide as the native host. Additionally, the activity of GccF produced by these systems is identical to that of the native producer with 2 nM being sufficient to inhibit the growth of L. plantarum ATCC 8014 by 50 %. Mutations have been introduced within the coding sequence of five of the cluster genes (gccA and gccC-F), confirming their roles in the production and maturation of GccF. One of the genes (gccE) remains an enigma, the C-terminal is a LytTR DNA binding domain and the N-terminal has no sequence homology to any other known protein. GccE is likely involved in controlling GccF production but is not a typical two component response regulator. Mutations in each domain have produced interesting results that indicate it could be involved in regulation but uses an unconventional signal. Details of these findings will be discussed in terms of current research.