The ice-nucleation protein (INP) from Pantoea ananatis was expressed in Escherichia coli. INP expression increased the freezing point of the E. coli culture by a few degrees. Deletion of FabH, an important enzyme in fatty acid biosynthesis, significantly inhibited the ice-nucleation activity. Increased unsaturated fatty acids in the fabH mutant cells decreased the ice-nucleation activity. Adding exogenous saturated fatty acids increased both E. coli fatty acid saturation and the ice-nucleation activity. In contrast, adding unsaturated fatty acids exhibited the opposite effects. Furthermore, an E. coli MG1655-fadR strain with high saturated fatty acids content was constructed, in which the INP activity was enhanced by about 17% compared with its activity in the wild-type MG1655 strain.