The goal of this study was to increase the production of long-chain fatty acids and to change the composition of fatty acids through the overexpression of genes involved in the fatty acid synthase (FAS) pathway and utilizing characteristics of a specific gene, namely, fabF. The four genes, fabB, fabG, fabZ, and fabI, are Escherichia coli homologues and function in the elongation cycle of fatty acid biosynthesis. FabB (fabB), an activator of FAS, is a beta-oxoacyl-ACP synthase, which catalyzes the addition of acyl-ACP to malonyl-ACP to generate beta-oxoacyl-ACP. FabF (fabF) participates at the same step as FabB in the elongation cycle and is structurally and functionally similar to FabB. Hence, we attempted to see if FabF was an activator of FAS, like FabB, with the rationale that these two enzymes have striking similarities. FabF exhibits thermal regulation in that enzyme activity increases at lower temperatures. To confirm its role as an activator of FAS, fabF was overexpressed solely or with other genes in the elongation cycle through biochemical engineering. The fabF recombinants were cultured at different temperatures, resulting in increased total and unsaturated fatty acid accumulation in all the recombinants, compared to wild type, at lower temperatures.