Expression of the Chlamydia psittaci major outer membrane protein in Escherichia coli: A model for translocation and vaccine development.

摘要

Chlamydia infections represent a serious ongoing health problem throughout the world. The major outer membrane protein (MOMP) of Chlamydia species is an abundant component of the chlamydial outer membrane and is the target of neutralizing antibodies in infected individuals. The entire omp1 gene encoding MOMP from Chlamydia psittaci strain GPIC has been cloned and expressed in Escherichia coli. A tightly regulated T7 promoter is used to control expression of the protein in E. coli. Upon induction of expression, the precursor MOMP (pre-MOMP) is synthesized in the cell. This is followed by the appearance of a lower molecular weight protein that comigrates with mature MOMP from chlamydial elementary bodies on both one-dimensional SDS-PAGE and two-dimensional gels. MOMP is not detected in surface labeling experiments using several MOMP-specific antibodies. In addition, MOMP expressed in E. coli is not accessible to protease treatment without prior permeabilization of the outer membrane. These data indicate that pre-MOMP is translocated to the periplasmic space and processed but is not surface-exposed in E. coli.; Biochemical and genetic techniques have been used to characterize the chlamydial homolog of Hsp60. Coexpression in E. coli of MOMP or a MOMP-PhoA fusion with Hsp60, or other chaperones, does not alter the level of MOMP translocation. The Hsp60 protein from C. psittaci was purified from isolated organisms. Hsp60 was investigated for the presence of intermolecular complexes with translocated chlamydial proteins. Chlamydial Hsp60 can function in trans to suppress a lethal temperature sensitive mutation of a cytoplasmic protein in E. coli.; Highly purified recombinant MOMP and Omp2, another cysteine-rich chlamydial outer membrane protein, were generated. These proteins were used in immunological studies of protective chlamydial antigens. Only MOMP could prime a protective antibody response in BALB/c mice. C3H/HeJ mice, a strain non-responsive to MOMP, failed to elicit protective antibodies using various combinations of recombinant MOMP and Omp2 as immunogens. An attenuated Salmonella typhimurium strain was used to deliver both MOMP and Omp2 by oral immunization but was unsuccessful in eliciting anti chlamydial antibodies.