Molecular cloning, expression, and sequence of the pilin gene from nontypeable Haemophilus influenzae M37.
Nontypeable Haemophilus influenzae M37 adheres to human buccal epithelial cells and exhibits mannose-resistant hemagglutination of human erythrocytes. An isogenic variant of this strain which was deficient in hemagglutination was isolated. A protein with an apparent molecular weight of 22,000 was present in the sodium dodecyl sulfate-polyacrylamide gel profile of sarcosyl-insoluble proteins from the hemagglutination-proficient strain but was absent from the profile of the isogenic hemagglutination-deficient variant. A monoclonal antibody which reacts with the hemagglutination-proficient isolate but not with the hemagglutination-deficient isolate has been characterized. This monoclonal antibody was employed in an affinity column for purification of the protein as well as to screen a genomic library for recombinant clones expressing the gene. Several clones which contained overlapping genomic fragments were identified by reaction with the monoclonal antibody. The gene for the 22-kDa protein was subcloned and sequenced. The gene for the type b pilin from H. influenzae type b strain MinnA was also cloned and sequenced. The DNA sequence of the strain MinnA gene was identical to that reported previously for two other type b strains. The DNA sequence of the strain M37 gene is 77% identical to that of the type b pilin gene, and the derived amino acid sequence is 68% identical to that of the type b pilin.
Coleman
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Edward Mallinckrodt Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri.
Grass
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Munson
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D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D003001:Cloning, Molecular; D036781:Fimbriae Proteins; D010861:Fimbriae, Bacterial; D015870:Gene Expression; D006193:Haemophilus influenzae / Q000235:genetics*; D006384:Hemagglutination; D051379:Mice; D008969:Molecular Sequence Data
D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins; D036781:Fimbriae Proteins