Cloning, characterization and sequence of a novel 59-kDa protein of Chlamydia trachomatis

Chlamydia trachomatis (Ct) serovar L2 DNA was partially digested with BamHI, ligated with plasmid vector pBR325 and used to transform Escherichia coli JMB83. Recombinant colonies were screened for their ability to synthesize chlamydial (chl) proteins by dot immunoblot and by in vitro transcription translation assays. A clone, B1, expressing a 59-kDa protein was further characterized, and the encoding gene was subcloned in the expression vector, pKK223-3, containing the tac promoter. Elevated levels of the 59-kDa protein were produced in E. coli in the presence of the lac inducer, IPTG. Sequencing identified one long open reading frame encoding a polypeptide of 59,075 Da (59 kDa). The partially purified 59-kDa protein was recognized by sera from patients with chl infections as shown in immunoblotting. In addition, the 59-kDa protein was located in the sarcosyl-soluble fraction of chl lysates. When used as a DNA probe in dot hybridization assays, the clone encoding the 59-kDa protein showed high homology to all serovars of Ct and four strains of Chlamydia psittaci. The cloned 59-kDa protein is neither related to the 60-kDa heat-shock protein found in many strains of bacteria, nor to the Cys-rich sarcosylinsoluble protein described in other studies of chlamydia.