Trichosanthin (TCS) is one of the single chain ribosome-inactivating proteins (RIPs). The crystals of the orthorhombic form of trichosanthin have been obtained from a citrate buffer (pH 5.4) with KCl as the precipitant. The crystal belongs to the space group P2(1)2(1)2(1) with a = 38.31, b = 76.22, c = 79.21 A. The structure was solved by molecular replacement method and refined using the programs XPLOR and PROLSQ to an R-factor of 0.191 for the reflections within the 6-1.88 A resolution range. The bond length and bond angle in the protein molecule have root-mean-square deviations from ideal value of 0.013 A and 3.3 degrees, respectively. The refined model includes 247 residues and 197 water molecules. The TCS molecule consists of two structural domains. The large domain contains six alpha-helices, a six-stranded sheet, and an antiparallel beta-sheet. The small domain has a largest alpha-helix, which shows a distinct bend. The possible active site of the molecule located on the cleft between two domains was proposed. In the active site Arg-163 and Glu-160, Glu-189 and Arg-122 form two ion pairs, Glu-189 and Gln-156 are hydrogen bonded to each other. Three water molecules are bonded to the residues in the active site region. The structures of TCS molecule and ricin A-chain (RTA) superimpose quite well, showing that the structures of the two protein molecules are homologous. Comparison of the structures of the TCS molecule in this orthorhombic crystal with that in the monoclinic crystal indicates that there are no essential differences of the structures between the two protein crystals.