SS-B/La is a conserved cellular phosphoprotein of 46/48 KD molecular weight which is the target antigen of autoantibodies in sera of patients with Sjögren's syndrome. Two relatively protease-resistant domains (X and Y) were defined in the SS-B antigen from HeLa cells. Human autoantibodies to SS-B were used as reagents. Domain X is a methionine-containing, non-phosphorylated 28 KD polypeptide and is resistant to partial digestion with six different proteases. Domain Y is a 23 KD polypeptide which contains little if any methionine, but all the detectable phosphorylated amino acids. These results demonstrate that there are at least two distinct antigenic epitopes on the 46/48 KD SS-B protein, each located on a separate structural domain. The asymmetric distribution of methionine and phosphorylated amino acid residues in SS-B show striking similarity to two reported domains of the adenovirus 72 KD DNA-binding protein, which also have separate methionine and phosphorylated amino acid distributions.