Organization of the AAA+ Adaptor Protein PspA Is an Oligomeric Ring

117

Vipp1 (vesicle-inducing protein in plastids 1) is found in Cyanobacteria and chloroplasts of photosynthetic eukaryotes where it is essential for the formation of the thylakoid membrane. Vipp1 is closely related to the phage shock protein A (PspA), a bacterial protein induced under diverse stress conditions. Vipp1 proteins differ from PspA by an additional C-terminal domain that is required for Vipp1 function in thylakoid biogenesis. We show here that in Cyanobacteria, green algae, and vascular plants, Vipp1 is part of a high molecular mass complex. The complex is formed by multiple copies of Vipp1, and complex formation involves interaction of the central ␣-helical domain that is common to Vipp1 as well as to PspA proteins. In chloroplasts of vascular plants, the Vipp1 complex can be visualized by green fluorescent protein fusion in discrete locations at the inner envelope. Green fluorescent protein fusion analysis furthermore revealed that complex formation is important for proper positioning of Vipp1 at the inner envelope of chloroplasts.Oxygenic photosynthesis is a specific feature of Cyanobacteria and chloroplasts of plants and algae. Characteristic of oxygenic photosynthesis is a specialized membrane system, the thylakoids, on which the components of the photosynthetic machinery are located (1). Despite its importance in the process of oxygenic photosynthesis, very little is known about the origin of the thylakoid membrane system in the ancestry of present day Cyanobacteria. Furthermore, proteins and other factors that are involved in the formation and maintenance of the thylakoid membrane in either Cyanobacteria or chloroplasts are not well defined.We have shown recently that in Arabidopsis as well as in Synechocystis the Vipp1 1 protein is essential for thylakoid biogenesis (2, 3). ⌬-vipp1 mutants, in which the expression of the vipp1 gene is greatly reduced, have lost their ability to build up a thylakoid membrane system and to perform oxygenic photosynthesis (2, 3). Interestingly, disruption of the vipp1 gene in Arabidopsis also abolished chloroplast vesicle transport, which has an alleged function in thylakoid formation in the chloroplasts of higher plants (2, 4). Vipp1 is closely related to PspA, a bacterial protein that is induced under distinct stress conditions, i.e. invasion by filamentous phages, inhibition of lipid biosynthesis, secretion defects, or the saturation of the twin arginine pathway (5-8). PspA and Vipp1 are distinguished by a C-terminal domain of about 20 -30 amino acids which is present in all Vipp1 proteins but lacking in PspA (3). It is presumed that this C-terminal extension is important for the role of Vipp1 in thylakoid biogenesis. Most Cyanobacteria possess genes coding for both PspA and Vipp1. Chloroplasts seem to have retained solely Vipp1, indicating that the function of PspA is no longer required in this organelle.So far, the exact function of PspA and Vipp1 remains elusive. In bacteria, PspA is part of a larger operon encoding four more proteins, PspB-PspE. PspA is induce...

Section: Discussionmentioning

confidence: 98%

Complex Formation of Vipp1 Depends on Its α-Helical PspA-like Domain

Aseeva

Ossenbühl

Eichacker

et al. 2004

117

“…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).…”

Section: Discussionmentioning

confidence: 99%

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

Collins

Frye

Kitmitto

et al. 2004

118

123

The bacterial pathogen Neisseria meningitidis expresses long, thin, retractile fibers (called type IV pili) from its cell surface and uses these adhesive structures to mediate primary attachment to epithelial cells during host colonization and invasion. PilQ is an outer membrane protein complex that is essential for the translocation of these pili across the outer membrane. Here, we present the structure of the PilQ complex determined by cryoelectron microscopy to 12 Å resolution. The dominant feature of the structure is a large central cavity, formed by four arm features that spiral upwards from a squared ring base and meet to form a prominent cap region. The cavity, running through the center of the complex, is continuous and is effectively sealed at both the top and bottom. Analysis of the complex using selforientation and by examination of two-dimensional crystals indicates a strong C4 rotational symmetry, with a much weaker C12 rotational symmetry, consistent with PilQ possessing true C4 symmetry with C12 quasisymmetry. We therefore suggest that the complex is a homododecamer, formed by association of 12 PilQ polypeptide chains into a tetramer of trimers. The structure of the PilQ complex, with its large and well defined central chamber, suggests that it may not function solely as a passive portal in the outer membrane, but could be actively involved in mediating pilus assembly or modification.

“…Notably, quasi-symmetry is a feature of membrane and membraneassociated protein complexes such as PilQ 2 The PspA AAAϩ adaptor protein assembly forms a C9-symmetric ring associated with the inner membrane (44). The molecular mass of the complex is ϳ1 MDa and suggests that each of the nine rotational units is a tetramer.…”

Section: Resultsmentioning

confidence: 99%

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

Beis

Collins

Ford

et al. 2004

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.