Envelope proteins

The major outer membrane protein: OmpA


The chlamydial outer membrane complex is composed primarily of three proteins; the major outer membrane protein (MOMP) and two Antigens_Proteomics.CysteineRichProteins, the outer membrane complex B protein (OmcB) and the outer membrane complex A protein (OmcA). The chlamydial major outer membrane protein, which by reason of its importance is the most extensively studied, was discovered in 1981 by three independent laboratories. The enabling technology was the relatively new technique of SDS - PAGE electrophoresis [lay reader: a method of sorting detergent-solubilised proteins in a polyacrylamide gel on the basis of their mobility in an electric field]. Hatch et al., 1981 described the presence of the MOMP in both the then known chlamydial species, C. psittaci and C. trachomatis.

Salari and Ward, 1981 [the author's lab in Southampton] described the molecular heterogeneity of the protein of molecular weight approximately 40 KDa in 14 of the 15 then known C. trachomatis serovars. They demonstrated the surface exposure of the protein by radio-iodination, suggested that it was immunodominant and the likely structural basis of the well known serotype variability of C. trachomatis.

Caldwell et al., 1981 working on _ C. trachomatis_ serovar L2, described an important method for the purification of the chlamydial outer membrane complex by its insolubility in the detergent sodium lauroyl sarcosinate. They too demonstrated the surface nature of the protein and performed a partial characterization. In their time these were seminal and complementary studies, with all three groups appreciating the potential importance of the protein.

MOMP and serological classification of C. trachomatis_

It soon became clear that MOMP was both structurally and immunologically the dominant protein in the chlamydial outer membrane complex. In artificial laboratory membranes MOMP functioned as a porin [or pore] protein [Bavoil, Ohlin & Schachter, 1984], though it was not entirely typical of known outer envelope proteins. Exposure to thiol reducing agents lead to a reduction in -S-S- intramolecular bonding of MOMP, opening up its pore structure for nutrient ingress and perhaps partly triggering the early differentiation of EBs after entry into the cell. The porin-like nature of MOMP was subsequently elegantly confirmed by the elegant electrical studies of native and recombinant MOMP in planar lipid bilayers [Wyllie _et al., 1998; 1999]. Like other porins, MOMP had a predominantly beta sheet content (62%) and was weakly anion selective and permeable to ATP, a key, energy rich molecule. These channels were modified by antibody [Wyllie et al., 1998]. Both polyclonal [Caldwell & Perry, 1982] and monoclonal [Zhang et al., 1987] antibodies to MOMP were capable of neutralizing chlamydial infectivity for tissue culture cells, though relatively large amounts were required. Monoclonal antibodies reactive with MOMP and raised against whole organisms showed type, subspecies and species - specific reactivities, with those against the type and subspecies specific regions of the protein most associated with neutralizing antibody [Zhang et al., 1987].

A major breakthrough came with the cloning, sequencing and expression of the omp_A gene encoding MOMP, a difficult task at the time, which was first achieved by Stephens _et al., 1995; 1996 for C. trachomatis serovar L2 and, subsequently, by Pickett et al., 1987. This revealed that ompA, exhibits extensive DNA sequence variation [unlike C. pneumoniae:]; the corollary of the molecular weight variation observed by Salari & Ward, 1981. In this respect it was quite different to C. pneumoniae where there is relatively little variability in MOMP [Carter et al., 1991].

Variation of C. trachomatis MOMP was primarily confined to four variable segments / domains (termed VS or VD 1 to VD 4) [Baehr _ et al_., 1998; Yuan et al., 1989] that contain subspecies- and serovar-specific antigenic determinants. The typing of C. trachomatis_ is based on the serological differentiation of antigenic epitopes on MOMP into 19 human _C. trachomatis serovars (A to K, Ba, Da, Ia, Ja, L1 to L3, and L2a). Based on amino acid similarity, these serovars have been placed into the following serogroups or classes: B class (B, Ba, D, Da, E, L1, L2, and L2a), C class (A, C, H, I, Ia, J, Ja, K, and L3), and intermediate class (F and G) [Yuan et al., 1989; Wang & Grayston, 1991; Wang et al., 1985].

Broadly speaking, type specific antibodies show specificities for one of these serovars only; sub-species specific antibodies for the class to a varying extent; species specific antibody for the whole C. trachomatis _species. The structural basis for these specificities were determined independently by Stephens _et al., 1988 and Conlan _ et al._, 1988. Initially, Stephens _ et al_., 1988 localised the serovar specific determinant of serovar L2 to a 14 amino acid peptide in VS 2 and the overlapping subspecies and species specific epitopes to a 16-amino acid peptide from VS 4. Conlan et al., 1988; 1989 introduced the PepscanŽ technique of Mario Geysen to chlamydial research and were able to map the serovar subspecies and species specific epitopes of C. trachomatis serovar L1 to single amino acid resolution. This is shown in Fig 1. They were also able to define the residues which were critical for binding [Conlan et al., 1989].

Fig 1.

Serovar specific Serovar Amino acid composition (single letter code)*
C. trachomatis C 141 TKTQSSSFNTAKL IP NT A 158
Sub-species and Species specific    
C. trachomatis C 290 LAEA ILDVTTL NPTIA G 306
Fig 1. Computer predicted serovar, subspecies and species specific epitopic regions of C. trachomatis MOMP. Underlined regions have an overall turn potential of p>1.5 x 10-4 . As a rough rule of thumb, surface exposed turn regions in protein folds tend to be associated with antigenic regions of proteins. Proteins consist of linear sequences of amino acids, which are here represented by a single letter. The red letters show critical binding residues of the serovar specific L1 epitope in VS 2. The green the L1 sub-species specific epitope and the yellow the species specific epitope in VS 4. Amino acids have been numbered (superscript) by reference to serovar C MOMP. For further information and an explanation of the single letter amino acid code, see Conlan, Clarke & Ward (1988) Molecular Microbiology 2, 673 - 679 from which this figure has been adapted.

The peptide approach was greatly extended by Zhong & Brunham, 1990; 1991; by Zhong et al., 1990; 1994 and by Batteiger 1996 & Batteiger _ et al_., 1996 to which the reader is referred. In particular, Batteiger 1996 in a study of genital serovars of _ C. trachomatis_ found that serovar specific epitopes associated with protection comprised all the central portion of VS1, (residues 70 to 77); the amino-terminal half of VS2, (residues 139 to 149); and the carboxyl-terminal third of VS4, (residues 305 to 315).

Clearly since MOMP is immunodominant and generates host antibodies which are protective, there was much interest in the question of whether MOMP might be a useful component of a defined chlamydial vaccine. Chlamydiae are difficult to bulk grow, making it difficult to produce significant quantities of native MOMP. There was therefore much interest as to whether the genetically engineered ompA gene, expressed in _ E. coli_ or some other amenable organism, might produce useful recombinant MOMP whose production could be scaled up for vaccine development. High level expression of chlamydial MOMP was first achieved in E. coli by Pickett et al., 1988, [see: Fig 2.]

Fig 2.

MOMP.gif (37195 bytes)

Fig 2. Transmission electron micrograph of dark amorphous 'lumps' of chlamydial major outer membrane protein (M) expressed in the gut bacterium Escherichia coli. Highly condensed intracellular membrane protein like this tends not to adopt a native shape. This can be a problem for vaccine development if a protective antibody is directed against shape rather than primary amino acid sequence, as is often the case. See: Pickett, Ward & Clarke (1988). Molecular Microbiology 2, 681 - 685.

Furthermore it was demonstrated_ that fragments of recombinant MOMP produced antibodies which bound to peptides corresponding to known neutralizing epitopes on MOMP, as well as to the surface of native elementary bodies [Conlan _et al., 1990]. However it is questionable whether non-native recombinant proteins like this produce adequately effective antibodies against the corresponding native protein. This is because the binding of antibody frequently depend on three dimensional shape as well as on the primary amino acid sequence. In general recombinant MOMP or peptides derived there from, despite early promise [Tuffrey _ et al_., 1990], have proved disappointing as components for a chlamydial vaccine.

[MEW] May 2002

NEXT: Antigens_Proteomics.IntraserovarRecombinationMOMP


Baehr, W., Zhang, Y. Z., Joseph, T., Su, H., Nano, F. E., Everett, K. D. and Caldwell, H. D. (1988). Mapping antigenic domains expressed by Chlamydia trachomatis major outer membrane protein genes. Proceedings of the National Academy of Science of the USA 85, 4000 - 4004. [Key paper at the time, comparing early MOMP sequences and coming up with some important conclusions]

Batteiger, B. E. (1996). The major outer membrane protein of a single Chlamydia trachomatis serovar can possess more than one serovar-specific epitope. Infection and Immunity 64, 542 - 547. Full article [Acrobat]

Batteiger, B. E., Lin, P. M., Jones, R. B. & van der Pol, B. J. (1996). Species-, serogroup-, and serovar-specific epitopes are juxtaposed in variable sequence region 4 of the major outer membrane proteins of some Chlamydia trachomatis serovars. Infection and Immunity 64, 2839 - 2841. Full article

Bavoil, P., Ohlin, A. & Schachter, J. (1984). Role of disulfide bonding in outer membrane structure and permeability in Chlamydia trachomatis. Infection and Immunity 44, 479 - 485. [Innovative study at the time whose results have largely been substantiated by Wyllie et al., 1998; 1999]

Caldwell, H. D., Kromhaut, J. and Schachter, J. (1981). Purification and partial characterization of the major outer membrane protein of Chlamydia trachomatis. Infection and Immunity 31, 111 - 116.

Caldwell, H. D. & Perry, L. J. (1982). Neutralization of Chlamydia trachomatis infectivity with antibodies to the major outer membrane protein. Infection and Immunity 38, 745 - 754. [Established that MOMP generates neutralizing antibody]

Carter, M. W., al-Mahdawi, S. A. H., Giles, I. G., Treharne, J. D., Ward, M. E. and Clarke, I. N. (1991). Nucleotide sequence and taxonomic value of the major outer membrane protein gene of Chlamydia pneumoniae IOL-207. Journal of General Microbiology 137, 465 - 475 [The initial ompA gene sequence for C. pneumoniae and first attempt at computerised classification of the Chlamydiaceae].

Conlan, J. W., Clarke, I. N. & Ward, M. E . (1988). Epitope mapping with solid phase peptides: identification of type-, subspecies-, species- and genus-reactive antibody binding domains on the major outer membrane protein of Chlamydia trachomatis. _Molecular Microbiology 2, 673 - 679. [High resolution mapping of epitopes on MOMP using combinatorial peptide synthesis. Described the critical binding residues of these epitopes].

Conlan, J. W., Ferris, S., Clarke, I. N. & Ward, M. E. (1990). Isolation of recombinant fragments of the major outer-membrane protein of Chlamydia trachomatis: their potential as subunit vaccines. Journal of General Microbiology 136, 2013 - 2020.

Conlan, J. W., Kajbaf, M., Clarke, I. N., Chantler, S. & Ward, M. E. (1989). The major outer membrane protein of Chlamydia trachomatis: critical binding site and conformation determine the specificity of antibody binding to viable chlamydiae. Molecular Microbiology 3, 311 - 318.

Hatch, T. P., Vance, D. W.Jr., Al-Hossainey, E. (1981). Identification of a major envelope protein in Chlamydia spp. Journal of Bacteriology 146, 426 - 431.

Hatch, T. (1999). Developmental biology, p. 29-68. In [Stephens, R. S. ed.], Chlamydia: intracellular biology, pathogenesis and immunity. ASM Press, Washington, D.C. [Good review]

Pickett, M. A., Ward, M. E. & Clarke, I. N. (1987). Complete nucleotide sequence of the major outer membrane protein gene from Chlamydia trachomatis serovar L1. FEMS Microbiology Letters 42, 185 - 190. [Permitted first omp_A sequence comparisons to be made]

Pickett, M. A., Ward, M. E. & Clarke, I. N. (1988). High-level expression and epitope localization of the major outer membrane protein of Chlamydia trachomatis serovar L1. _Molecular Microbiology 2, 681 - 685.

Salari, S. H. & Ward, M. E. (1981). Polypeptide composition of Chlamydia trachomatis. Journal of General Microbiology 123, 197 - 205. [First evidence molecular heterogeneity of MOMP associated with serovars]

Stephens, R. S., Kuo, C-C., Newport, G. & Agabian, N. (1985). Molecular cloning and expression of Chlamydia trachomatis major outer membrane protein antigens in Escherichia coli. Infection and Immunity 47, 713 - 718.

Stephens, R. S., Mullenbach, G., Sanchez-Pescador, R. & Agabian, N. (1986). Sequence analysis of the major outer membrane protein gene from Chlamydia trachomatis serovar L2. Journal of Bacteriology 168, 1277 - 1282. [The key first sequence].

Stephens, R. S., Wagar, E. A. & Schoolnik, G. K. (1988). High resolution mapping of serovar-specific and common antigenic determinants of the major outer membrane protein of Chlamydia trachomatis. Journal of Experimental Medicine 167, 817 - 831.

Tuffrey, M., Alexander, F., Conlan, W., Woods, C., Ward, M. E. (1992). Heterotypic protection of mice against chlamydial salpingitis and colonization of the lower genital tract with a human serovar F isolate of Chlamydia trachomatis by prior immunization with recombinant serovar L1 major outer-membrane protein. Journal of General Microbiology 138, 1707 - 1715.

Wang, S. P., and J. T. Grayston. 1991. Three new serovars of Chlamydia trachomatis: Da, Ia, and L2a. Journal of Infectious Diseases 163, 403 - 405. [When is a serovar a serovar, or just a variant?]

Wang, S. P., Kuo, C-C., Barnes, R. C., Stephens, R. S. & Grayston, S. T. (1985). Immunotyping of Chlamydia trachomatis with monoclonal antibodies. Journal of Infectious Diseases 152, 791 - 800.

Wyllie, S., Ashley, R. H., Longbottom, D. & Herring, A. J. (1998). The major outer membrane protein of Chlamydia psittaci functions as a porin-like ion channel. Infection and Immunity 66, 5202 - 5207. [Elegant study confirming and extending Bavoil et al., 1984].

Wyllie S., Longbottom, D., Herring, A. J. & Ashley, R. H. (1999). Single channel analysis of recombinant major outer membrane protein porins from Chlamydia psittaci and Chlamydia pneumoniae. FEBS Letters 445, 192 - 196.

Yuan, Y., Y. Zhang, X., Watkins, N. G. & Caldwell, H. D. (1989). Nucleotide and deduced amino acid sequences for the four variable domains of the major outer membrane proteins of the 15 Chlamydia trachomatis serovars. Infection and Immunity 57, 1040 - 1049.

Zhong, G., Berry, J. & Brunham, R. C. (1994). Antibody recognition of a neutralization epitope on the major outer membrane protein of Chlamydia trachomatis. Infection and Immunity 62, 1576 - 1583.

Zhong, G. M. & Brunham, R. C. (1990). Immunoaccessible peptide sequences of the major outer membrane protein from Chlamydia trachomatis serovar C. Infection and Immunity 58, 3438 - 3441.

Zhong, G. M. & Brunham, R. C. (1991) Antigenic determinants of the chlamydial major outer membrane protein resolved at a single amino acid level. Infection and Immunity 59, 1141 - 1147.

Zhong, G. M., Reid, R. E. & Brunham, R. C. (1990). Mapping antigenic sites on the major outer membrane protein of Chlamydia trachomatis with synthetic peptides. Infection and Immunity 58, 1450 - 1455.

NEXT: Antigens_Proteomics.IntraserovarRecombinationMOMP in MOMP

Edit | Attach | Watch | Print version | History: r4 < r3 < r2 < r1 | Backlinks | Raw View | Raw edit | More topic actions
Topic revision: r4 - 2011-02-04 - MeWard
This site is powered by the TWiki collaboration platform Powered by PerlCopyright © 2008-2017 by the contributing authors. All material on this collaboration platform is the property of the contributing authors.
Ideas, requests, problems regarding TWiki? Send feedback