Shigellosis is a major public health problem in many developing countries. Antibiotic therapy can reduce the severity of the dysentery and prevent potentially lethal complication. However, owing to the increased resistance to most of the widely used and inexpensive antibiotics, there is an urgent need for new antibacterial agents, particularly those that act on novel targets. Chorismate synthase (CS) is a key enzyme in the shikimic acid pathway, which is essential for the synthesis of aromatic amino acids in bacteria. As an anti-bacterial drug target, CS has been well validated. A homology model of Shigella-CS with the flavin mononucleotide (FMN) binding was constructed using the crystal structure of CS from other species. The substrate 5-enolpyruvylshikimate 3-phosphate (EPSP) was subsequently docked into the active site based on previous theoretical studies. Molecular dynamics (MD) was used to refine the starting ternary model. The model was well conserved during the 1.8 ns MD simulation with the equilibrium root mean square deviation (RMSD) value of 3.5 angstrom. The substrate binding energy was calculated and the electrostatic energy was found to be the most important term for binding. Decomposition of binding energies revealed that R129, R125, R327, R134 and R48 are important residues involved in substrate binding, which is useful for further site-directed mutagenesis experiments. In the absence of crystal structure, our study provides an early insight into the structure of CS from Shigella flexneri and its binding to the substrate and cofactor, thus facilitating the inhibitor design.