Curcumin (CUR) is a natural polyphenol with broad-spectrum antimicrobial efficacy, however, its use has been limited by low water solubility and poor stability. The aim of this study was to develop CUR nanocrystals (NCs) to improve its water solubility and stability, and investigate the effect of surface charge on its antibacterial activity. CUR NCs were prepared with different ratios of Pluronic F127 (F127) and hexadecyl trimethyl ammonium bromide (CTAB) as stabilizers to achieve tunable surface charge. The mean particle sizes of CUR NCs were around 150 nm, and zeta potentials were varied from −16.7 to 36.5 mV. Solubility and stability of CUR were significantly enhanced by processing into NCs. In vitro dissolution test showed that the dissolution rate of CUR NC was significantly higher than free CUR. Compared with free CUR, CUR NCs displayed low cytotoxicity against normal human kidney-2 cells. Moreover, comprehensive experiments (MIC, MBC, zone of inhibition and colony counting method) all demonstrate that CUR NCs with high positive surface charge exhibited excellent antibacterial activity than free CUR against E. coli and S. aureus, and SEM images showed that CUR NCs could destroy the surface of the bacteria cell to enhance the antibacterial activity. The overall results suggest that the CUR NC may be a promising antibacterial agent for pharmaceutical application.