The H4 histone of the protozoan Tetrahymena pyriformis, as obtained previously (Nomoto, M. & Iwai, K. (1982) J. Biochem. 91, 719-723), was completely sequenced; the total sequence reported preliminarily (Hayashi, H., Nomoto, M., & Iwai, K. (1980) Proc. Jpn. Acad. 56B, 579-584) is one of the two variant sequences determined here. The intact H4 was directly sequenced by automated Edman degradation from the N-terminal through residue 92. Sequence determination was further performed with tryptic peptides and peptic peptides, both covering the whole sequences. Thus, the complete sequences of two variants were determined; both consist of a total of 102 amino acid residues, have identical compositions, have the same molecular weights of 11,228 in the unmodified form, and are partially acetylated at four lysine residues from the N-terminal. The sequences differ in two positions from each other (-Lys-Arg-/-Arg-Lys- at residues 19 and 20, 7 : 3 mol/mol), and in 22 or 20 positions from the human spleen H4 sequence. The implications of these results for the structure-function relationship of this histone species and also for the phylogeny of protozoa are discussed.