Nisin is a peptide that possesses potent antibacterial properties. This study evaluated the antibacterial activity of a nisin-doped adhesive against Streptococcus mutans, as well as its degree of conversion and microtensile bond strength (μTBS) to dentin. Nisin was added to the adhesive Adper Single Bond 2 (3M ESPE), resulting in four groups: Control Group (Single Bond 2); Group 1% (1 wt% nisin-incorporated), Group 3% (3 wt% nisin-incorporated) and Group 5% (5 wt% nisin-incorporated). Antibacterial activity against S. mutans was evaluated using colony-forming unit counts (CFU). The degree of conversion was tested using FTIR. Forty human teeth were restored for μTBS evaluation. Data were statistically analyzed with ANOVA and Tukey tests at α = 0.05. The nisin-doped adhesives, for all concentrations, exhibited a significant inhibition of the growth of S. mutans (p < 0.05); Incorporation of 5% and 3% nisin decreased the degree of conversion of the adhesive (p < 0.05). The μTBS (in MPa): Control Group—38.3 ± 2.3A, Group 1%—35.6 ± 2.1A, Group 3%—27.1 ± 1.6B and Group 5%—22.3 ± 1.0C. Nisin-doped adhesives exerted a bactericidal effect on S. mutans. The μTBS and degree of conversion of adhesive were not affected after incorporation of 1% nisin.