Three-dimensional Structure of Wza, the Protein Required for Translocation of Group 1 Capsular Polysaccharide across the Outer Membrane of Escherichia coli

Abstract

Wza is a highly conserved multimeric outer membrane protein complex required for the surface expression of the serotype K30 group 1 capsular polysaccharide in Escherichia coli. Here we present the first three-dimensional structure of this type of polysaccharide exporter at a 15.5-Å resolution obtained using single particle averaging on a dataset of cryo-negatively stained protein.Previous structural studies on purified Wza have revealed a homo-oligomeric ring structure that is most probably composed of eight subunits. Symmetry analysis of the three-dimensional structure combined with biochemical two-and three-dimensional crystallographic data strongly suggest that Wza is an octameric complex with a C4 quasi-rotational symmetry and is organized as a tetramer of dimeric subunits. Wza is best described as a stack of two 4-Å high rings with differing diameters providing a mushroom-like aspect from the side. The larger ring has a distinctive square shape with a diameter of 115 Å, whereas the smaller is almost circular with a diameter of 90 Å. In the center of the complex and enclosed by the four symmetrical arms is a small elliptical cagelike cavity of ϳ40 Å in diameter. The central cavity is effectively sealed at the top and bottom of the complex but has small inter-arm holes when viewed from the side. We discuss the structure of this complex and implications in the surface translocation of cell-surface polysaccharide.

Roles of the Structural Symbiosis Polysaccharide (syp) Genes in Host Colonization, Biofilm Formation, and Polysaccharide Biosynthesis in Vibrio fischeri

“…However, SypC differs in size from other Wza proteins (full-length SypC is 703 amino acids in length, whereas Wza proteins from E. coli and Vibrio vulnificus are only 379 and 410 amino acids, respectively), suggesting that it may also carry out an additional function. Both the E. coli and V. fischeri proteins have or appear to have a signal peptide that would direct them across the inner membrane, but neither protein has the ␤-barrel structure typical of outer membrane proteins (1). We found that the putative signal sequence was important for SypC function, as the deletion of the N terminus abrogated the function of SypC.…”

Structure of the Neisseria meningitidis Outer Membrane PilQ Secretin Complex at 12 Å Resolution

“…Such quasi-symmetry is well characterized for viral capsids, but has been described relatively infrequently outside this group of structures. We note, however, that quasi-symmetry is a feature of some other membrane protein structures, including the EmrE homodimer (36), the BetP trimer (37), the polysaccharide K antigen transporter Wza (38), and also the membrane-associated "adaptor" protein PspA (39). There is also evidence that the quaternary structure of the PapC usher involves quasi-symmetry (40).…”

Pivotal Roles of the Outer Membrane Polysaccharide Export and Polysaccharide Copolymerase Protein Families in Export of Extracellular Polysaccharides in Gram-Negative Bacteria

“…The images suggested Wza to be a channel-forming protein (28,79). Subsequent single-particle analyses of Wza oligomers visualized by cryo-negative-staining EM (15.5-Å resolution), revealed a novel hollow, barrel-like structure (dimensions, 90 by 90 by 100 Å) (10). The unique features of Wza were reinforced with a 2.3-Å-resolution crystal structure (26).…”