PSEUDOMONAS-AERUGINOSA PA-I LECTIN GENE MOLECULAR ANALYSIS AND EXPRESSION IN ESCHERICHIA-COLI

This communication describes a Pseudomonas aeruginosa DNA fragment (cloned in lambda gt11) which contains the structural gene coding for the galactophilic PA-I lectin (pa-1L, 369 bp) and an additional downstream 237 bp sequence. This DNA is relatively rich in G + C (54%), and exhibits a strong codon preference biased for XXC and also for XXG. The Shine-Dalgarno site of the gene is preceded by an adjacent ATATAT sequence resembling the - 10 sequence of the Escherichia coli promoter. The stop codons are followed by a stem and loop structure - typical of the rho-independent transcriptional stop element. This lambda gt11-cloned DNA was expressed in E. coli Y1090 cells. The resulting cell lysates exhibited a galactose-specific hemagglutination and a protein with electrophoretic mobility similar to that of the native PA-I, which were both absent from E. coli lysates infected with ovalbumin gene-bearing bacteriophages. The recombinant PA-I, purified by gel filtration and affinity chromatography, was shown to be a galactophilic hemagglutinin resembling the native lectin in molecular weight and selective reactivity with rabbit anti native PA-I serum. These results are important for development of a safe Pseudomonas aeruginosa vaccine using recombinant DNA techniques, thus avoiding contamination with toxic products of this bacterium.