Characterization and partial purification of the human receptor for the heat‐stable enterotoxin

The receptor for the Escherichia coli heat‐stable enterotoxin has been characterized and partially purified from the T84 human colonic cell line. Using a novel mutant heat‐stable enterotoxin peptide as a radioligand (the C‐terminal tyrosine residue is replaced by phenylalanine in the mutant), a single class of high‐affinity receptor sites was detected in T84 cells, with a K d of 0.1 nM, similar in affinity to the receptor described in human intestinal tissue. The receptor was solubilised from T84 cell membranes and affinity cross‐linking of the solubilised preparation indicated that a single species of M r 160000 served as the receptor. Freshly solubilised preparations of the receptor retained heat‐stable enterotoxin‐activable guanylyl cyclase activity. Purification of the receptor was achieved through sequential affinity chromatography on GTP–epoxy‐Sepharose and wheat‐germ‐agglutinin columns resulting in purification of the receptor by 3000 fold. The heat‐stable enterotoxin‐binding characteristics of the receptor were unchanged during the purification and silver staining of the purified receptor preparation indicated a band of M r 160000, which was specifically cross‐linked to the 125 I‐labeled mutant peptide. The purified receptor retained guanylyl cyclase activity, but the activity was not stimulated on addition of human heat‐stable enterotoxin, suggesting that accessory structural factors may be involved in the activation of the guanylyl cyclase/receptor.