Isolation, affinity purification, and identification of piglet small intestine mucosa receptor for enterotoxigenic Escherichia coli k88ac+fimbriae

An affinity chromatography technique was utilized to isolate and purify the receptors of Escherichia coli K88ac(+) fimbriae from the mucus of the smal l intestines of newborn piglets. Purified K88ac+ fimbriae were covalently i mmobilized onto a beaded agarose matrix (Sepharose 4B). The immobilized fim briae were used for the affinity purification of the K88ac+ receptors. Only two major proteins were tightly and specifically bound to the immobilized fimbriae after the column containing bound receptor was washed exhaustively with a buffer containing a high concentration of salt and a detergent. The receptors were eluted as a single component at a low pH. The isolated prot eins were then subjected to enzyme-linked immunosorbent assay, sodium dodec yl sulfate-polyacrylamide gel electrophoresis, and Western blot (immunoblot ) analyses. The two proteins were of high purity, were responsible for near ly all of the fimbrial binding capacity of the crude mucus, and had molecul ar masses of 26 and 41 kDa. The method for isolation of E. coli binding pro teins is simple and yields purified intestinal receptors in a single chroma tographic run. The intestinal mucus of different piglets has different prop ortions of the two receptor proteins.