Recombinant glycans on an S-layer self-assembly protein: a new dimension for nanopatterned biomaterials.
Crucial biological phenomena are mediated through carbohydrates that are displayed in a defined manner and interact with molecular scale precision. We lay the groundwork for the integration of recombinant carbohydrates into a "biomolecular construction kit" for the design of new biomaterials, by utilizing the self-assembly system of the crystalline cell surface (S)-layer protein SgsE of Geobacillus stearothermophilus NRS 2004/3a. SgsE is a naturally O-glycosylated protein, with intrinsic properties that allow it to function as a nanopatterned matrix for the periodic display of glycans. By using a combined carbohydrate/protein engineering approach, two types of S-layer neoglycoproteins are produced in Escherichia coli. Based on the identification of a suitable periplasmic targeting system for the SgsE self-assembly protein as a cellular prerequisite for protein glycosylation, and on engineering of one of the natural protein O-glycosylation sites into a target for N-glycosylation, the heptasaccharide from the AcrA protein of Campylobacter jejuni and the O7 polysaccharide of E. coli are co- or post-translationally transferred to the S-layer protein by the action of the oligosaccharyltransferase PglB. The degree of glycosylation of the S-layer neoglycoproteins after purification from the periplasmic fraction reaches completeness. Electron microscopy reveals that recombinant glycosylation is fully compatible with the S-layer protein self-assembly system. Tailor-made ("functional") nanopatterned, self-assembling neoglycoproteins may open up new strategies for influencing and controlling complex biological systems with potential applications in the areas of biomimetics, drug targeting, vaccine design, or diagnostics.
Steiner
Kerstin
K
University of Natural Resources and Applied Life Sciences, Center for NanoBiotechnology, Gregor-Mendel-Strasse 33, A-1180 Wien, Austria.
Hanreich
Angelika
A
Kainz
Birgit
B
Hitchen
Paul G
PG
Dell
Anne
A
Messner
Paul
P
Schäffer
Christina
C
D000942:Antigens, Bacterial / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D001672:Biocompatible Materials / Q000378:metabolism*; D016123:Campylobacter jejuni / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000648:ultrastructure; D005818:Genetic Engineering; D001411:Geobacillus stearothermophilus / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000648:ultrastructure; D006031:Glycosylation; D008956:Models, Chemical; D053758:Nanoparticles; D019897:Periplasm / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism*; D011135:Polysaccharides, Bacterial / Q000648:ultrastructure; D021381:Protein Transport; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D001672:Biocompatible Materials; D006023:Glycoproteins; D011134:Polysaccharides; D011135:Polysaccharides, Bacterial