Cloning, expression and sequence analysis of the gene encoding the 120kD surface‐exposed protein of Rickettsia rickettsii

Abstract

Rickettsia rickettsii (R strain) genomic DNA was partially digested and cloned into the lambda expression vector gt11 generating a genomic clone bank. Transformant plaques were screened with antisera generated against the 120 kiloDalton protein of R. rickettsii to detect those phage expressing the recombinant protein. The gene encoding the 120 kD protein was localized to a 4.3 kilobase SphI-BamHI fragment from the recombinant phage and subcloned into pUC18 and pUC19. Full-length expression of the recombinant protein was achieved with both orientations. The gene and flanking regions were sequenced. The p120 gene consists of 3900 base pairs coding for 1300 amino acids. A distinguishable promoter region was not identified, although there are several 5' sequences that resemble classical prokaryotic promoters. Downstream of the termination codon for this gene lies a 726 base pair open reading frame on the opposite strand with the potential to encode a protein of approximately 27 kD. The identity of this putative gene product is unknown. The two open reading frames are separated by a 106 base pair intergenic region that consists of a stretch of dyad symmetry resembling rho-independent transcriptional terminators.

Cross‐Reactivity of Rickettsia japonica and Rickettsia typhi Demonstrated by Immunofluorescence and Western Immunoblotting

“…Since IgG antibodies produced for protein in an early stage of infection and for carbohydrates are known to possess a low affinity, such antibodies carry a high specificity that causes cross-reaction with a very similar or identical antigenic structure (7,16). Gilmore et al (10,11) demonstrated the cross-reactivity of rOmp B between SFG and TG and confirmed the ompB gene homology by Southern blot analysis using the cloned DNA of R. rickettsii ompB. The sequencing of the R. japonica ompB gene revealed the existence in rOmp B of a portion consisting of some 50 putative amino acid residues identical to those in R. typhi (Yan, Y., Uchiyama, T., and Uchida, T., manuscript in preparation).…”

Genotypic identification and phylogenetic analysis of the spotted fever group rickettsiae by pulsed-field gel electrophoresis

“…Five genes have currently been cloned and sequenced: the genes encoding citrate synthase (62), ADP/ATP translocase (61), and three surface proteins of 17 (3), 120 (27), and 190 (2) kDa in R nickettsii. We have located three of them on our profiles.…”

Molecular analysis of a gene encoding a serum-resistance-associated 76 kDa surface antigen of Haemophilus somnus

“…An example of the former situation was found for the 120 kDa Rickettsia rickettsii OMP (Gilmore et al, 1989(Gilmore et al, , 1991, which contains enough DNA in the ORF to encode a deduced 168 kDa protein.…”