Expression of the Chlamydia trachomatis major outer membrane protein-encoding gene in Escherichia coli: role of the 3' end in mRNA stability.
The major outer membrane protein (MOMP)-encoding gene (omp1) of Chlamydia trachomatis has been cloned into Escherichia coli and partially sequenced. This recombinant gene expresses a full-length 40-kDa product, which is recognized by a monoclonal antibody directed against the species-specific epitope of MOMP. The recombinant omp1 is expressed in either insertion orientation, indicating that it utilizes its own promoter system. The endogenous omp1 promoter possesses a relatively low activity despite the high level of MOMP expression. Deletion of a 520-bp fragment at the 3' end encoding 39 amino acids (aa) at the C terminus and the remainder of the noncoding region leads to a significant decrease in mRNA stability and loss of protein synthesis. When the MOMP-encoding plasmid was introduced into E. coli minicells, it expressed 40- and 43-kDa proteins; however, inhibition of post-translational processing by ethanol revealed only a 43-kDa protein. These data indicate that the unprocessed omp1 gene product contains a 22-aa leader sequence which is cleaved during translocation to the outer membrane, to yield a processed 40-kDa protein. The recombinant MOMP was localized to the outer membrane E. coli fraction, comparable to the location of the native C. trachomatis protein.
Kaul
R
R
Department of Pediatrics, University of Alberta, Edmonton, Canada.
Duncan
M J
MJ
Guest
J
J
Wenman
W M
WM
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D002692:Chlamydia trachomatis / Q000235:genetics*; D002872:Chromosome Deletion; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics*; D015183:Restriction Mapping
D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D012333:RNA, Messenger