A systematic and extensive conformational search has been performed to characterize the gas-phase tryptophan structures. A total of 648 unique trial structures were generated by allowing for all combinations of internal single-bond rotamers and were optimized at the DFT/B3LYP/6-311G* level of theory. A total of 45 local minima conformers were found. Further optimization of the 45 conformers with B3LYP and MP2/6-311++G** did not produce meaningful structural change, and accurate geometries, dipole moments, rotational constants, harmonic frequencies, and relative energies were then determined. Combined with statistical mechanics principles, the conformational distributions of gas-phase tryptophan at different temperatures are shown. The results clearly support the conclusion drawn by Compagnon et al. that only one dominant isomer existed in the molecular beam at 85 K and add further evidence that the supersonic jet expansion or embedding helium droplets did not produce an equilibrium distribution.