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X L Rudner, Z Zheng, R S Berk, R T Irvin, L D Hazlett; Corneal epithelial glycoproteins exhibit Pseudomonas aeruginosa pilus binding activity.. Invest. Ophthalmol. Vis. Sci. 1992;33(7):2185-2193.
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Adherence of Pseudomonas aeruginosa to the cornea is a requisite step in the pathogenesis of bacteria-induced corneal disease. P. aeruginosa is capable of attaching to host epithelial cells by its pili, but there is little information regarding the epithelial receptors of this adhesin in the cornea. Using nitro-cellulose blotting of polyacrylamide gels of solubilized adult mouse corneal epithelium, four major proteins (molecular weights: 38, 42, 57, and 66 kD) and several minor proteins were identified that bound purified pili from strain PAK and its hyperpiliated mutant PAK/PR1. These proteins were identified by immunoblotting either with pilus-specific monoclonal antibodies, XLR-3 and PK 3B, or using peptide PAK 128-144 (OX). The glycosylated nature of the proteins was determined using similar gel electrophoresis of corneal epithelial proteins, blotting onto nitrocellulose, and staining the blots with lectins conjugated to either horseradish peroxidase or alkaline phosphatase. All four major pilus-binding proteins were stained with concanavalin A lectin (mannose and glucose) and either wheat germ agglutinin lectin (WGA, specific for sialic acid and N-acetylglucosamine) or succinylated WGA lectin (only N-acetylglucosamine). Staining for peanut agglutinin lectin (galactose beta(1-3) N-acetylgalactosamine) was seen for the 42-, 57-, and 66-kD proteins. The importance of the carbohydrate portions of these corneal proteins in pili binding was confirmed by preincubation of corneal epithelial blots with periodate or pili with sialic acid, both of which abolished the pili binding. These studies indicate that corneal epithelial pilus-binding proteins are glycoproteins in nature and that sialic acid may be a constituent of these pilus-specific receptors in the adult mouse corneal epithelium.
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