The
nisin leader is believed to be crucial for
nisin biosynthesis. Here, by using a construct completely lacking the
leader peptide, we show that an up to fivefold-dehydrated leaderless prenisin can be obtained, as judged by MALDI-TOF MS, and that some of these species are biologically active, thus suggesting that at least three
lanthionine rings are present. Notably, by expressing the
leader peptide in trans together with the leaderless prenisin, we were able to increase the
dehydration/cyclization efficiency of both NisB and NisC, but still with limited efficiency until the fifth dehydratable residue (Thr13) was processed, thereby enabling three rings to form. This, for the first time, demonstrates that 1) the leader is not absolutely necessary for the
dehydration reaction of class I
lantibiotics to occur in vivo; 2) the leader acts in trans in vivo; 3) the leader increases the efficiency of modification. Based on previous work and our current study, a model for the interactions of NisB and NisC with prenisin is proposed, in which the leader induces a more active conformation and/or productive complex formation of the biosynthetic machinery, and, when covalently bound, is involved in increasing the efficiency of
dehydration to the C-terminal end of the prenisin substrate molecule.