Expression of a plasmid gene of Chlamydia trachomatis encoding a novel 28 kDa antigen.
Plasmid pCT is present in essentially all isolates of Chlamydia trachomatis and may encode factors important for survival in the natural environment. However, no pCT-associated phenotype has been described so far. With the purpose of investigating the possibility of a role of pCT in C. trachomatis pathogenicity we examined the expression of an ORF (ORF3), potentially encoding a 28 kDa polypeptide (pgp3). Analysis of RNA extracted from chlamydia-infected Vero cells detected ORF3-specific transcripts, from 20 h post-infection onwards, mainly as discrete RNA species of 1390 nucleotides comprising the downstream ORF4 sequence. ORF3 DNA was cloned and expressed in Escherichia coli as a 39 kDa fusion protein (MS2/pgp3). Antibodies raised against purified MS2/pgp3, specifically recognized a 28 kDa protein on Western blots of protein from purified chlamydial elementary bodies (EBs). The same antibodies detected chlamydial inclusions in methanol-fixed infected cells by immunofluorescence. Western blot analysis of EBs extracted with 2% Sarkosyl, showed that a large proportion of the 28 kDa antigen is associated with the detergent-insoluble ('membrane') fraction. Antibodies recognizing pgp3 epitopes were detected in sera from patients with chlamydial infections, but not in sero-negative control sera. The finding support the hypothesis that pCT may provide a function related to chlamydial cell physiology.
Comanducci
M
M
Immunobiological Research Institute Siena (IRIS), Italy.
Cevenini
R
R
Moroni
A
A
Giuliani
M M
MM
Ricci
S
S
Scarlato
V
V
Ratti
G
G
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D000942:Antigens, Bacterial; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D001483:Base Sequence; D002690:Chlamydia Infections / Q000276:immunology; D002692:Chlamydia trachomatis / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics*; D006801:Humans; D008565:Membrane Proteins / Q000302:isolation & purification; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D010957:Plasmids / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics*; D014709:Vero Cells
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D008565:Membrane Proteins; D012333:RNA, Messenger; C082169:pgp3 protein, Chlamydia