Nisin, the only antimicrobial peptide that has GRAS (generally considered as safe) status from both WHO and FDA, has the potential to become an important natural antibiotic as bacteria become more resistant to traditional antibiotics. Many studies have focused on how modification of nisin affects antimicrobial activity. In this study, nisin and chitosan were crosslinked using genipin to modify the N-terminal of nisin. The antimicrobial experiments were carried out on Staphylococcus aureus ATCC 6538. The results showed that nisin still retained antimicrobial activity after modification. When the crosslinking ratio of nisin to chitosan increased to 200: 1, MIC decreased from 48 μg/ml to 40 μg/ml. Less chitosan increased local number density and antimicrobial activity of nisin. When the local number density was high, adjacent peptides could cooperatively penetrate into the lipid membrane. The results of this study may help to better understand the antimicrobial mechanism and provide suggestions for the further modification of nisin.