The Polygonal Cell Shape and Surface Protein Layer of Anaerobic Methane-Oxidizing Methylomirabilis lanthanidiphila Bacteria.

Lavinia Gambelli,Christoph A Diebolder,Mike S M Jetten,Bertram Daum,Andreas Engel,Wouter Versantvoort,Laura Van Niftrik,Rob Mesman,Martin Pabst,Wiel Evers

doi:10.3389/fmicb.2021.766527

Abstract

Methylomirabilis bacteria perform anaerobic methane oxidation coupled to nitrite reduction via an intra-aerobic pathway, producing carbon dioxide and dinitrogen gas. These diderm bacteria possess an unusual polygonal cell shape with sharp ridges that run along the cell body. Previously, a putative surface protein layer (S-layer) was observed as the outermost cell layer of these bacteria. We hypothesized that this S-layer is the determining factor for their polygonal cell shape. Therefore, we enriched the S-layer from M. lanthanidiphila cells and through LC-MS/MS identified a 31 kDa candidate S-layer protein, mela_00855, which had no homology to any other known protein. Antibodies were generated against a synthesized peptide derived from the mela_00855 protein sequence and used in immunogold localization to verify its identity and location. Both on thin sections of M. lanthanidiphila cells and in negative-stained enriched S-layer patches, the immunogold localization identified mela_00855 as the S-layer protein. Using electron cryo-tomography and sub-tomogram averaging of S-layer patches, we observed that the S-layer has a hexagonal symmetry. Cryo-tomography of whole cells showed that the S-layer and the outer membrane, but not the peptidoglycan layer and the cytoplasmic membrane, exhibited the polygonal shape. Moreover, the S-layer consisted of multiple rigid sheets that partially overlapped, most likely giving rise to the unique polygonal cell shape. These characteristics make the S-layer of M. lanthanidiphila a distinctive and intriguing case to study.

Highlights

Methylomirabilis S-Layer and Cell Shape attributed to “Candidatus Methylomirabilis oxyfera,” a bacterium belonging to the NC10 phylum (Raghoebarsing et al, 2006)
We find that the surface protein layer (S-layer) has a hexagonal symmetry and is composed of a single repeating protein unit identified as mela_00855
The distance between the S-layer and the outer membrane remained constant, indicating that the S-layer proteins are anchored to the outer membrane, but not to the cytoplasmic membrane that did not have a polygonal shape

Summary

Introduction

Methylomirabilis S-Layer and Cell Shape attributed to “Candidatus Methylomirabilis oxyfera,” a bacterium belonging to the NC10 phylum (Raghoebarsing et al, 2006). The outermost layer of the cell envelope contains multiple longitudinal ridges that end in a cap-like structure at the cell poles This outermost layer of M. oxyfera was proposed to be a surface protein layer (S-layer), about 8 nm thick, of oblique or square symmetry (Wu et al, 2012). Since S-layers, have been described to promote and maintain the cell shape of bacteria, it was postulated that the S-layer of M. oxyfera is the responsible factor for its polygonal shape (Wu et al, 2012). Maintaining a rod or filamentous shape (or any shape other than a sphere) requires the cell to counterbalance the surface tension and the cell osmotic pressure, which would force the cell into a sphere (Young, 2004) For this reason, the evolution of a robust cell envelope and a cytoskeleton are paramount (van Teeseling et al, 2017). It has become apparent that S-layer proteins show a low degree of conservation on the level of the primary sequence (Sleytr et al, 2014)

Methods

Results

Conclusion