Nisin A is a potent antimicrobial with potential as an alternative to traditional antibiotics, and a number of genetically modified variants have been created that target clinically relevant pathogens. In addition to antimicrobial activity, nisin autoregulates its own production via a signal transduction pathway, a property that has been exploited in a protein expression system termed the nisin-controlled gene expression (NICE) system. Although NICE has become one of the most popular protein expression systems, one drawback is that the inducer peptide, nisin A, also has inhibitory activity. It has already been demonstrated that the N-terminal region of nisin A contributes to antimicrobial activity and signal transduction properties; therefore, we conducted bioengineering of nisin at positions Pro9 and Gly10 within ring B to produce a bank of variants that could potentially be used as alternative induction peptides. One variant, designated nisin M, has threonines at positions 9 and 10 and retains induction capacity comparable to that of wild-type nisin A, while most of the antimicrobial activity is abolished. Further analysis confirmed that nisin M produces a mix of peptides as a result of different degrees of dehydration of the two threonines. We show that nisin M exhibits potential as a more suitable alternative to nisin A for the expression of proteins that may be difficult to express or for production of proteins in strains that are sensitive to wild-type nisin. Moreover, it may address the increasing demand by industry for optimization of peptide fermentations to increase yields or production rates. This study describes the generation of a nisin variant with superior characteristics for use in the NICE protein expression system. The variant, termed nisin M, retains an induction capacity comparable to that of wild-type nisin A but exhibits significantly reduced antimicrobial activity and can therefore be used at concentrations that are normally toxic to the expression host.