Identifying assembly-inhibiting and assembly-tolerant sites in the SbsB S-layer protein from Geobacillus stearothermophilus.

Surface layer (S-layer) proteins self-assemble into two-dimensional crystalline lattices that cover the cell wall of all archaea and many bacteria. We have generated assembly-negative protein variants of high solubility that will facilitate high-resolution structure determination. Assembly-negative versions of the S-layer protein SbsB from Geobacillus stearothermophilus PV72/p2 were obtained using an insertion mutagenesis screen.

Genetic engineering of the S-layer protein SbpA of Lysinibacillus sphaericus CCM 2177 for the generation of functionalized nanoarrays.

The mesophilic organism Lysinibacillus sphaericus CCM 2177 produces the surface (S)-layer protein SbpA, which after secretion completely covers the cell surface with a crystalline array exhibiting square lattice symmetry. Because of its excellent in vitro recrystallization properties on solid supports, SbpA represents a suitable candidate for genetically engineering to create a versatile self-assembly system for the development of a molecular construction kit for nanobiotechnological applications. The first goal of this study was to investigate the surface location of 3 different C-terminal amino acid positions within the S-layer lattice formed by SbpA.

Recognition imaging and highly ordered molecular templating of bacterial S-layer nanoarrays containing affinity-tags.

Functional nanoarrays were fabricated using the chimeric bacterial cell surface layer (S-layer) protein rSbpA fused with the affinity tag Strep-tagII and characterized using various atomic force microscopy (AFM) techniques in aqueous environment. The accessibility of Strep-tagII was verified by single-molecule force spectroscopy studies employing Strep-Tactin as specific ligand. Simultaneous topography and recognition imaging (TREC) of the nanoarray yielded high resolution maps of the Strep-tagll binding sites with a positional accuracy of 1.

Surface location of individual residues of SlpA provides insight into the Lactobacillus brevis S-layer.

Bacterial surface layer (S-layer) proteins are excellent candidates for in vivo and in vitro nanobiotechnological applications because of their ability to self-assemble into two-dimensional lattices that form the outermost layer of many Eubacteria and most Archaea species. Despite this potential, knowledge about their molecular architecture is limited. In this study, we investigated SlpA, the S-layer protein of the potentially probiotic bacterium Lactobacillus brevis ATCC 8287 by cysteine-scanning mutagenesis and chemical modification.