Diverse S-layer anchoring mechanisms create an epiplasmic space in prokaryotic cell envelopes.

Surface layers (S-layers) are paracrystalline protein lattices that form the outermost boundary of many archaeal and bacterial cells. Interest in S-layer biology has grown rapidly in recent years, driven by advances in structural and bioinformatic studies that have revealed new insights into lattice organization and function. In contrast, the mechanisms that anchor these lattices to the underlying cell envelope during growth, division, and cellular morphogenesis remain comparatively understudied. Because anchoring is dictated by the chemistry and architecture of the underlying cell envelope, we propose that S-layer anchoring strategies provide valuable insights into prokaryotic evolution. Based on recent experimental work and bioinformatic predictions, we summarize the major anchoring mechanisms found across archaea and bacteria. Our analysis reveals a recurring feature of S-layers: the arrangement of lattice-forming and envelope-proximal anchoring regions, frequently connected by linkers, generates an enclosed space near the underlying cell envelope, with an enigmatic biological role. We discuss possible contributions of this enclosed 'epiplasmic' space to mechanical scaffolding, chemical compartmentalization, and substrate harvesting.