Characterization of P40, a cytadhesin of Mycoplasma agalactiae.
An immunodominant protein, P40, of Mycoplasma agalactiae was analyzed genetically and functionally. The gene encoding P40 was cloned from type strain PG2, sequenced, submitted to point mutagenesis in order to convert mycoplasma-specific TGA(Trp) codon to the universal TGG(Trp) codon, and subsequently expressed in Escherichia coli. Nucleotide sequence-derived amino acid sequence comparisons revealed a similarity of P40 to the adhesin P50 of Mycoplasma hominis and to protein P89 of Spiroplasma citri, which is expected to be involved in adhesion. The amino acid sequence of P40 revealed a recognition site for a signal peptidase and strong antigenic and hydrophilic motifs in the C-terminal domain. Triton X-114 phase partitioning confirmed that P40 is a membrane protein. Fab fragments of antibodies directed against recombinant purified P40 significantly inhibited adherence of M. agalactiae strains PG2 to lamb joint synovial cells LSM 192. Sera taken sequentially from sheep infected with PG2 revealed that P40 induced a strong and persistent immune response that gave strong signals on immunoblots containing recombinant P40 even 3 months after infection. The gene encoding P40 was present in a single copy in all of the 26 field strains of M. agalactiae analyzed and was not detected in closely related mycoplasma species. P40 was expressed as a protein with an apparent molecular mass of 37 kDa on sodium dodecyl sulfate-acrylamide gels by all M. agalactiae strains except for serotype C strains, which showed nonsense mutations in their p40 genes.
Fleury
Bénédicte
B
Unité Mixte de Recherche ENVT-INRA 959, Ecole Nationale Vétérinaire de Toulouse, F-31076 Toulouse Cedex 3, France.
Bergonier
Dominique
D
Berthelot
Xavier
X
Peterhans
Ernst
E
Frey
Joachim
J
Vilei
Edy M
EM
D018829:Adhesins, Bacterial / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics; D001422:Bacterial Adhesion; D001483:Base Sequence; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D005260:Female; D005798:Genes, Bacterial; D006041:Goats; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009174:Mycoplasma / Q000235:genetics* / Q000276:immunology* / Q000472:pathogenicity; D009175:Mycoplasma Infections / Q000382:microbiology / Q000662:veterinary; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D012756:Sheep; D012757:Sheep Diseases / Q000382:microbiology; D013045:Species Specificity
D018829:Adhesins, Bacterial; D000942:Antigens, Bacterial; D004269:DNA, Bacterial; D011994:Recombinant Proteins