The Major Outer Membrane Protein of Chlamydia psittaci Functions as a Porin-Like Ion Channel

Abstract

The major outer membrane protein (MOMP) of Chlamydiaspecies shares several biochemical properties with classical porin proteins. Secondary structure analysis by circular dichroism now reveals that MOMP purified from Chlamydia psittaci has a predominantly β-sheet content (62%), which is also typical of bacterial porins. Can MOMP form functional ion channels? To directly test the “porin channel” hypothesis at the molecular level, the MOMP was reconstituted into planar lipid bilayers, where it gave rise to multibarreled channels, probably trimers, which were modified by an anti-MOMP monoclonal antibody. These observations are consistent with the well-characterized homo-oligomeric nature of MOMP previously revealed by biochemical analysis and with the triple-barreled behavior of other porins. MOMP channels were weakly anion selective (P Cl/P K ∼ 2) and permeable to ATP. They may therefore be a route by whichChlamydia can take advantage of host nucleoside triphosphates and explain why some anti-MOMP antibodies neutralize infection. These findings have broad implications on the search for an effective chlamydial vaccine to control the significant human and animal diseases caused by these organisms.

Single channel analysis of recombinant major outer membrane protein porins from Chlamydia psittaci and Chlamydia pneumoniae

“…We recently reported the puri¢cation, structural analysis and functional reconstitution of native OEA C. psittaci MOMP [6]. Circular dichroism (CD) indicated that MOMP has a L-sheet-rich, porin-like secondary structure and the reconstitution of MOMP into planar lipid bilayers gave rise to porin-like ion channels [6]. This work extended the liposome swelling studies of Bavoil et al [7] and added fresh insights into MOMP structure/function relationships.…”

“…The addition of similar volumes of OG/DTT alone had no e¡ect. Channel incorporation appeared to be autocatalytic, as previously described for native C. psittaci MOMP [6] and so after the ¢rst channels appeared the cis chamber was perfused with fresh 50 mM KCl in an attempt to limit the channel content. We counted the number of discrete unit conductances in 21 independent experiments with recombinant C. psittaci MOMP and as observed with the native protein [6], channels tended to appear in groups of three (Fig.…”

“…Whereas the conductances of native [6] and recombinant C. psittaci MOMPs were the same over a range of KCl activities, those of recombinant C. pneumoniae MOMP were signi¢cantly higher, as summarised in Table 1. There was, however, a signi¢cant di¡erence in relative anion versus cation selectivity between native and recombinant C. psittaci MOMPs.…”

“…The native C. psittaci MOMP was obtained as previously described [6]. To obtain recombinant proteins, the pET constructs described above were transformed into expression host BL21 (DE3) and protein expression was induced as described previously [13].…”

“…Recombinant MOMPs were incorporated into 0.3 mm diameter planar lipid bilayers cast from decane suspensions of diphytanoylphosphatidylcholine using standard reconstitution and recording methods, as previously described for native C. psittaci MOMP [6]. Control experiments were carried out using E. coli BL21(DE3) outer membrane proteins (Omps) which were also solubilised in OG/DTT and reconstituted in exactly the same way as inclusion body proteins.…”