A New Addition to the Cell Plan of Anammox Bacteria: "Candidatus Kuenenia stuttgartiensis" Has a Protein Surface Layer as the Outermost Layer of the Cell

Abstract

bAnammox bacteria perform anaerobic ammonium oxidation (anammox) and have a unique compartmentalized cell consisting of three membrane-bound compartments (from inside outwards): the anammoxosome, riboplasm, and paryphoplasm. The cell envelope of anammox bacteria has been proposed to deviate from typical bacterial cell envelopes by lacking both peptidoglycan and a typical outer membrane. However, the composition of the anammox cell envelope is presently unknown. Here, we investigated the outermost layer of the anammox cell and identified a proteinaceous surface layer (S-layer) (a crystalline array of protein subunits) as the outermost component of the cell envelope of the anammox bacterium "Candidatus Kuenenia stuttgartiensis." This is the first description of an S-layer in the phylum of the Planctomycetes and a new addition to the cell plan of anammox bacteria. This S-layer showed hexagonal symmetry with a unit cell consisting of six protein subunits. The enrichment of the S-layer from the cell led to a 160-kDa candidate protein, Kustd1514, which has no homology to any known protein. This protein is present in a glycosylated form. Antibodies were generated against the glycoprotein and used for immunogold localization. The antiserum localized Kustd1514 to the S-layer and thus verified that this protein forms the "Ca. Kuenenia stuttgartiensis" S-layer.

Structural Studies of Planctomycete Gemmata obscuriglobus Support Cell Compartmentalisation in a Bacterium

“…This makes the G.obscuriglobus cell surface similar to those of some Archaea [28] and to those of Gram-positive bacteria where a periplasm appears to exist despite the absence of a Gram-negative wall outer membrane [29], as well as perhaps to another well-studied compartmentalized planctomycete, Kuenenia suttgartiensis , where an S-layer protein wall overlies an outermost membrane [30]. Such Gram-negative bacteria as Caulobacter crescentus also have an outermost proteinaceous layer [31] but it is attached to a lipopolysaccharide-containing outer membrane and in addition they possess a typical peptidoglycan layer between the outer and inner membranes, and conventional bacterial cell division system [32].…”

Global and Targeted Lipid Analysis of Gemmata obscuriglobus Reveals the Presence of Lipopolysaccharide, a Signature of the Classical Gram-Negative Outer Membrane

“…The proteinaceous barrier was previously suggested to substitute for other classical Gram-negative cell features, such as cell surface protection more typically provided by O antigen (20), a rigid structural framework in place of peptidoglycan (13), and possibly as an attachment platform for degradative enzymes, as in a related species, Rhodopirellula baltica (21). The proteinaceous exterior of G. obscuriglobus has not been shown to constitute a surface layer (S-layer), typically composed of only one or two proteins in a lattice configuration, like that found in some other bacteria, including archaea and the planctomycete "Candidatus Kuenenia stuttgartiensis" (13,22,23). The original cell wall analysis of G. obscuriglobus by Stackebrandt et al (18) found the cell wall composition to include many amino acids, and the ability of the G. obscuriglobus cell to maintain its integrity was demonstrated by the finding that cell sacculi retained their native cell morphology after 10% SDS treatment (18), thus exhibiting resistance to SDS that is more like that of cells that have a peptidoglycan-containing murein sacculus (21).…”

Immunogold Localization of Key Metabolic Enzymes in the Anammoxosome and on the Tubule-Like Structures of Kuenenia stuttgartiensis

“…Adapted from reference 30. (35), outer membrane, peptidoglycan-containing periplasm (36), cytoplasmic membrane, cytoplasm, anammoxosome membrane, anammoxosome that contains tubule-like structures (white arrowheads), and electron-dense, iron-rich particles (black arrowheads). Note that not all tubule-like structures visible in the micrograph were visualized in the schematic overview.…”