ELONGATION-FACTOR TS OF CHLAMYDIA-TRACHOMATIS - STRUCTURE OF THE GENEAND PROPERTIES OF THE PROTEIN

A putative structural gene cluster containing four open reading frames (ORFs) located downstream of the omp1 gene of Chlamydia trachomatis m ouse pneumonitis (MoPn) was cloned and sequenced, A GenBank survey ind icated that the identified cluster is similar to the rpsB-tsf-pyrH(smb A)-frr region of Escherichia coli. The second ORF was 846 bp encoding a 282-amino-acid polypeptide with a calculated M-r 30,824. Alignment o f this deduced protein sequence and E. coli elongation factor Ts (EF-T s, product of tsf) demonstrated 34% identity and an additional 14% sim ilarity. The putative chlamydial tsf gene was expressed in E. coli as a nonfusion protein and as a 6x His-tagged fusion protein. By SDS-PAGE analysis, the molecular weights of the nonfusion recombinant protein and a protein of chlamydial elementary bodies (EBs), which was recogni zed by monoclonal antibodies derived from the nonfusion recombinant pr otein, are 34 kDa. The purified recombinant 6x His-tagged fusion prote in increased the rate of GDP exchange with both Chlamydia and E. coli elongation factor Tu (EF-Tu). These data show that the second gene of the identified cluster is tsf. Unlike EF-Ts from any other species, it s activity was comparable to that of E. coli EF-Ts in exchange reactio n with E. coli EF-Tu. (C) 1997 Academic Press.