Bacterial heme was produced from a genetic engineered Escherichia coli via porphyrin pathway and it was useful as an iron resource for animal feed. The amount of the E. coli synthesized heme, however, was only few milligram in a culture broth and it was not enough for industrial applications. To analyze heme biosynthetic pathways, an engineered E. coli artificially overexpressing ALA synthase (hemA from Rhodobacter sphaeroides) and pantothenate kinase (coaA gene from self geneome) was constructed as a bacterial heme producing strain, and both the transcription levels of pathway genes and the intermediates concentrations were determined from batch and continuous cultures. Transcription levels of the pathway genes were not significantly changed among the tested conditions. Intracellular intermediate concentrations indicated aminolevulinic acid (ALA) and coenzyme A (CoA) were enhanced by the hemA-coaA co-expressions. Intracellular coproporphyrinogen I and protoporphyrin IX accumulation suggested that the bottleneck steps in heme biosynthetic pathway could be the spontaneous conversion of HMB to coproporphyrinogen I and limited conversion of protoporphyrin IX to heme, respectively. Strategy to increase the conversion of ALA to heme is discussed based on the results.