ISOLATION AND CHARACTERIZATION OF PSEUDOMONAS-PSEUDOMALLEI FLAGELLIN PROTEINS

Flagellin proteins from several different strains of Pseudomonas pseud omallei have been isolated and purified to homogeneity by mechanical s hearing and differential centrifugation techniques. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis yielded flagellin monomer protein hands with an estimated M(r) of 43,400. No lipopolysac charide contamination of the purified protein preparations was detecta ble by silver staining of flagellin displayed on polyacrylamide gels a nd by Western immunoblotting with P. pseudomallei antilipopolysacchari de monoclonal antibody. NH2-terminal amino acid sequence analysis of t he flagellin protein of P. pseudomallei 319a revealed significant homo logy with flagellins from Proteus mirabilis, Bordetella bronchiseptica , and Pseudomonas aeruginosa PAK. Rabbit polyclonal antiserum raised a gainst the 319a flagellin protein reacted with 64 of 65 P. pseudomalle i strains tested. The polyclonal antiserum proved effective in inhibit ing the motility of these organisms in motility agar plates. Passive i mmunization studies demonstrated that 319a flagellin-specific antiseru m was capable of protecting diabetic rats from challenge with a hetero logous P. pseudomallei strain.