Protease digestion of colonic mucin. Evidence for the existence of two immunochemically distinct mucins.

Colonic mucin was prepared by phenol-water partition extraction of th colonic mucosa, followed by ethanol precipitation of the water-soluble material, molecular sieve chromatography, and reductive sodium dodecyl sulfate-disc gel electrophoresis in an agarose-polyacrylamide gradient gel. Although this material as observed to be relatively homogeneous by physical criteria, it was shown to contain at least two components by immunodiffusion. Molecular sieve, ion exchange, adsorption, lectin affinity, and electrophorectic techniques failed to separate the two components. However, by protease digestion employing either 1% w/w pronase or papain, followed by molecular sieve chromatography, we were able to separate at least the portions of the molecules containing the immunodeterminants. Physicochemical and immunologic properties of both components were characteristic of mucins. Mucin A (pronase digest fraction 1 from molecular sieve chromatography) was enriched in threonine, proline, and sialic acid, while mucin B (pronase digest fraction 3 from molecular sieve chromatography) was enriched in serine, alanine, and fucose. Mucin A and mucin b were shown to be immunologically distinct. The evidence suggests that the protease-digested mucin fractions originate from two different mucins, one a sialomucin, the other a fucomucin; however, the alternative explanation, that these two mucin fragments may originate from a single mucin molecule, cannot be ruled out at present.