Didemnin B, a highly active depsipeptide isolated from a Caribbean tunicate, crystallizes from chloroform/benzene in the orthorhombic space group C2221, with cell parameters a = 14.990 +/- 0.003 A, b = 22.574 +/- 0.004 A, c = 41.112 +/- 0.009 A, V = 13911.7 A3 at 138 K and a calculated density of 1.143 g/cm3 based on C57H89N7O15, 1.5C6H6.H2O and eight formula units per cell. The overall agreement factor R = 0.052 for 7699 reflections, 20 theta max = 150 degrees, Cu K-alpha radiation. The structure determination revealed that didemnin B contains an isostatine residue instead of a statine residue. The conformation of the 23-membered depsipeptide ring is stabilized by one transannular hydrogen bond. The ring does not show the antiparallel beta-pleated-sheet structure but, instead, has a fold in the shape of a bent figure-eight. The linear peptide moiety, containing N-methylleucine and lactylproline, forms a beta (II)-bend and is folded back toward the cyclic backbone, giving the overall molecule a globular character. Comparison with the structure of cyclosporin A shows distinct stereochemical differences between the two molecules. It is suggested that didemnin B and cyclosporin A are unlikely to have a common receptor binding site.