Crystallization and preliminary crystallographic studies of MOMP (major outer membrane protein) fromCampylobacter jejuni

Abstract

Campylobacter jejuni is the leading bacterial cause of human enteritis linked to ingestion of contaminated food or water. MOMP, the major outer membrane protein from these Gram-negative bacteria, belongs to the porin family. In order to determine the three-dimensional structure of this protein and to elucidate the underlying molecular mechanisms, the MOMP from C. jejuni strain 85H has been purified and crystallized by vapour diffusion. Two crystal forms were characterized for this membrane protein. X-ray diffraction data were collected to a resolution of 3.1 A using a synchrotron-radiation source from the orthorhombic crystal form, which belonged to space group P2(1)2(1)2 with unit-cell parameters a = 170.1, b = 101.9, c = 104.9 A. With a trimer in the asymmetric unit, the solvent content is 64% (V(M) = 3.4 A Da(-1)). The other form exhibits trigonal symmetry (space group R3) with hexagonal unit-cell parameters a = b = 94.2, c = 161.2 A, but diffracts X-rays poorly to about 4 A with significant anisotropy.

MOMP from Campylobacter jejuni Is a Trimer of 18-Stranded β-Barrel Monomers with a Ca 2+ Ion Bound at the Constriction Zone

“…To allay concerns about the “nativeness” of this recombinant structure arising from the difference in the lipid environment during expression, we also determined the structure of MOMP purified directly from Campylobacter and we found it to be essentially identical. Crystals, but no structure of MOMP from C. jejuni , have been previously reported [18]. The lower purity of the MOMP obtained directly from Campylobacter highlighted the key advantage of E. coli expression.…”

Infection-induced antibodies against the major outer membrane protein ofCampylobacter jejunimainly recognize conformational epitopes

“…At present, there are no effective vaccines available for prevention of Campylobacter colonization or infection in chickens and humans. The Campylobacter MOMP is characterized as a bbarrel transmembrane protein and exhibits three conformational forms including the folded monomer ($35 kDa), the denatured monomer (40-48 kDa), and the native trimer (120-140 kDa) (De et al, 2000;Zhang et al, 2000;Labesse et al, 2001;Bolla et al, 2004). The Campylobacter MOMP is characterized as a bbarrel transmembrane protein and exhibits three conformational forms including the folded monomer ($35 kDa), the denatured monomer (40-48 kDa), and the native trimer (120-140 kDa) (De et al, 2000;Zhang et al, 2000;Labesse et al, 2001;Bolla et al, 2004).…”

“…As determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), C. jejuni has an abundant OMP named MOMP (De et al, 2000;Zhang et al, 2000). The Campylobacter MOMP is characterized as a bbarrel transmembrane protein and exhibits three conformational forms including the folded monomer ($35 kDa), the denatured monomer (40-48 kDa), and the native trimer (120-140 kDa) (De et al, 2000;Zhang et al, 2000;Labesse et al, 2001;Bolla et al, 2004). Both the trimer and monomer of C. jejuni MOMP have pore-forming activities (De et al, 2000).…”

“…For designing vaccines for human use, recombinant MOMP (rMOMP) free of lipooligosaccharide is preferred, because the lipooligosaccharide antigen in Campylobacter can potentially trigger autoimmunity related to the Guillain-Barré syndrome (Nachamkin et al, 1998;Yuki, 2005). Although the sequence and structural features of Campylobacter MOMP have been well characterized in earlier studies (De et al, 2000;Zhang et al, 2000;Bolla et al, 2004), the nature of antibody responses to Campylobacter MOMP induced during natural infection is unknown and needs to be examined to facilitate the design of MOMP-based vaccines. In addition, a practical method that can effectively refold the rMOMP into its native state is required for vaccine preparation.…”

Properties and Phylogeny of 76 Families of Bacterial and Eukaryotic Organellar Outer Membrane Pore-Forming Proteins

“…Cluster II includes families that fall into two subclusters, each containing three families. Subcluster A includes the Cyanobacterial Trimeric OMPP (CBP) Family (1.B.23) [ 67 ], found in many bacterial phyla, the OMP50 OMPP (Omp50) Family (1.B.60) [ 88 ], represented in several Gram-negative bacterial phyla (these two families are more closely related) and the Putative Bacterial OMPP (PBP) Family with no functionally characterized members (1.B.62). The second subcluster in cluster II, subcluster IIB, includes the Short Chain Amide and Urea OMPP (SAP) Family (1.B.16), the Putative β-Barrel OMPP-4 (BBP4) Family (1.B.67) of unknown function, and the Pseudomonas OprP OMPP (POP) Family (1.B.5) of anion-selective OMPPs [ 89 , 90 ].…”