Characterization of the first planctomycetal outer membrane protein identifies a channel in the outer membrane of the anammox bacterium Kuenenia stuttgartiensis

• Published in: Biochimica et biophysica acta. Biomembranes Vol. 1860; no. 3; pp. 767 - 776
• Main Authors: van Teeseling, Muriel C.F., Benz, Roland, de Almeida, Naomi M., Jetten, Mike S.M., Mesman, Rob J., van Niftrik, Laura
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.03.2018
• Subjects: Ammonium Compounds - metabolism; Anammox bacteria; Bacterial Outer Membrane Proteins - isolation & purification; Bacterial Outer Membrane Proteins - metabolism; Cations - metabolism; Cell Membrane - ultrastructure; Cell Wall - ultrastructure; Gram-Negative Bacteria - ultrastructure; Immunohistochemistry; Ion Channels - isolation & purification; Ion Channels - metabolism; Ion Transport; Kuenenia stuttgartiensis; Kustd1878; Lipid Bilayers; Outer membrane protein; Planctomycetales - chemistry; Planctomycetales - metabolism; Planctomycetales - ultrastructure; Planctomycetes; Potassium - metabolism; Potassium Channels - isolation & purification; Potassium Channels - metabolism; Kuenenia stuttgartiensis; Kustd1878; Planctomycetes; Anammox bacteria; Outer membrane protein
• ISSN: 0005-2736; 1879-2642
• DOI: 10.1016/j.bbamem.2017.12.020

Planctomycetes are a bacterial phylum known for their complex intracellular compartmentalization. While most Planctomycetes have two compartments, the anaerobic ammonium oxidizing (anammox) bacteria contain three membrane-enclosed compartments. In contrast to a long-standing consensus, recent insights suggested the outermost Planctomycete membrane to be similar to a Gram-negative outer membrane (OM). One characteristic component that differentiates OMs from cytoplasmic membranes (CMs) is the presence of outer membrane proteins (OMPs) featuring a β-barrel structure that facilitates passage of molecules through the OM. Although proteomic and genomic evidence suggested the presence of OMPs in several Planctomycetes, no experimental verification existed of the pore-forming function and localization of these proteins in the outermost membrane of these exceptional microorganisms. Here, we show via lipid bilayer assays that at least two typical OMP-like channel-forming proteins are present in membrane preparations of the anammox bacterium Kuenenia stuttgartiensis. One of these channel-forming proteins, the highly abundant putative OMP Kustd1878, was purified to homogeneity. Analysis of the channel characteristics via lipid bilayer assays showed that Kustd1878 forms a moderately cation-selective channel with a high current noise and an average single-channel conductance of about 170–190pS in 1M KCl. Antibodies were raised against the purified protein and immunogold localization indicated Kustd1878 to be present in the outermost membrane. Therefore, this work clearly demonstrates the presence of OMPs in anammox Planctomycetes and thus firmly adds to the emerging view that Planctomycetes have a Gram-negative cell envelope. [Display omitted] •Kuenenia stuttgartiensis membranes harbour two major pore-forming proteins.•Kustd1878 is a cation-selective pore with a conductance of 170–190pS in 1M KCl.•Immunogold localization shows the presence of Kustd1878 in the outermost membrane.•This OMP further evidences that Planctomycetes have a Gram-negative cell envelope.