The interaction between membrane proteins and cytoplasmic structural proteins is thought to be one mechanism for maintaining the spatial order of proteins within functional domains on the plasma membrane1–4. Such interactions have been characterized extensively in the human erythrocyte5–7, where a dense, cytoplasmic matrix of proteins comprised mainly of spectrin and actin8, is attached through a linker protein, ankyrin, to the anion transporter (Band 3) 9,10. In several nonerythroid cell types, including neurons11–13, exocrine cells14 and polarized epithelial cells15,16 homologues of ankyrin and spectrin (fodrin) 11,12,15,17–23 are localized in specific membrane domains. Although these results suggest a functional linkage between ankyrin and fodrin and integral membrane proteins in the maintenance of membrane domains in nonerythroid cells, there has been little direct evidence of specific molecular interactions. Using a direct biological and chemical approach, we show here that ankyrin binds to the ubiquitous (Na+ + K+)ATPase, which has an asymmetrical distribution in polarized cells14, 24–30.