Latest SCI publications
Research project (§ 26 & § 27)
Duration : 2016-10-01 - 2021-09-30
Archaea are one of the oldest life-forms existing on Earth. These unicellular organisms are often adapted to extreme habitats. Since the cell envelope of many archaea consists only of a very thin layer of fat (lipid membrane), into which an outermost crystalline protein layer is anchored, the question arises how Nature can accomplish this high resistance to extreme environmental conditions. The present project will study the reassembly of cell envelopes of archaea using previously isolated biological components, i.e., lipids and proteins. It aims to clarify the question how the self-organization of etherlipids and surface layer proteins proceeds in detail and which anchoring strategies are available for the formation of an artificial cell membrane. In addition, the question is addressed which properties of the biomolecules themselves and their assembly into macroscopic cell envelopes cause this amazing resistance to the extreme habitat conditions. Hence, selected archaea strains will be bred in the bioreactor and subsequently the basic building blocks will be isolated from the biomass. In addition, the surface layer proteins can also be genetically produced by host cells. This is a new approach, which has previously not attempted by another research group. By the application of Nature’s construction principle, the cell envelope structure of archaea will be reconstructed layer by layer. Each step will be tracked and analyzed using modern microscopial and surfaces-sensitive techniques. The results of this project will provide valuable insights into the isolation and in particular the self-assembly of cell envelope components. This knowledge can be applied to produce surface coatings with very specific properties. Further application are biomimetic membrane systems for studying the cell walls of archaea. The latter may also serve as model systems into which membrane-active peptides and membrane proteins can be incorporated and systematically investigated.
Shedding light on the pyruvylation of cell wall polymers – an ancestral reaction for protein cell surface display in bacteria
Research project (§ 26 & § 27)
Duration : 2019-01-01 - 2020-12-31
Several bacteria from almost all phyla are covered with a two-dimensional crystalline array of self-assembling (glyco)proteins, termed cell surface (S)-layer. This S layer has nanometer-scale periodicity and, thus, provides the basis for envisaged applications in nanotechnology and biomedicine (1). However, without detailed knowledge of how the anchoring of the S-layer to the underlying peptidoglycan cell wall of bacteria is elaborated. Driven by the research on the pathogen Bacillus anthracis and the model organism Paenibacillus alvei CCM2051T, most knowledge is available on the situation in Gram-positive bacteria, where so-called surface layer homology (SLH) domains serve as cell wall targeting modules for S-layer proteins. SLH domains, in turn, interact with a species-specific, peptidoglycan-bound secondary cell wall polymer (SCWP) that serves as a cell wall ligand (2-4). While it is obvious that this protein cell surface strategy holds promises for therapeutic intervention, a mechanistic understaniding of the underlying principle is still missing. These SCWPs fall into the category of non-classical SCWPs, since structurally different form the well known teichoic acids (3). The structure of the P. alvei SCWP was fully elucidated in our laboratory to be a polymer composed of eleven [→3)-β-D-ManpNAc-(1→4)-β-D-GlcpNAc-(1→] disaccharide repeats where every D-β-ManNAc residue is modified with a 4,6-linked pyruvate ketal (2) contributing to the anionic character of this SCWP. Importantly, the pyruvylation of D-β-ManNAc, which is also present at terminal position of the B. anthracis SCWP (4), is regarded an indispensable and ancestral epitope in this protein cell surface display mechanism (5-7). The functional coupling of SLH domain containing proteins (SLH proteins) and SCWP pyruvylation is substantiated by the finding that several SLH protein syntheisizing bacteria with a pyruvate containing cell wall have an ortholog of the CsaB enzyme predicted to catalyze the transfer of pyruvate ketal to β-D-ManNAc (5). Considering the predictably wide-spread occurrence of this protein cell surface display mechanism in both pathogenic and non-pathogenic bacteria it is surprising how little is known about the biosynthesis of pyruvylated SCWPs and the involved enzymes. This study is designed to shed light on the pyruvyltransferase CsaB of P. alvei, predicted to catalyze a key step in the biosynthesis of the P. alvei SCWP which s the basis for the cell surface display of a specific class of proteins (SLH proteins). This includes the determination of the enzymes's substrate specificity, kinetics and the identification amino acids involved in substrate binding. Further, interaction of CsaB with other enzymes form the P. avlei SCWP biosynthesis gene locus will be investigated, thereby contributing to our general understanding of how SCWPs are biosynthesized.
Research project (§ 26 & § 27)
Duration : 2019-01-01 - 2021-12-31
Carbon nano tubes (CNTs) are cylindrical nano structures made of carbon atoms. Due to their outstanding mechanical and electrical properties, and thermal conductivity, they are already used as additives in various novel materials. Recently, CNTs have also been considered for several medical applications due to their small diameters and ability to penetrate cells and tissues. However, since CNTs are chemically inert and insoluble in water, they have to be chemically functionalized or coated with biomolecules to carry payloads or interact with the environment. Proteins bound to the surface of CNTs are preferred because they provide a better biocompatibility and offer functional groups for binding additional molecules. Nevertheless, their arrangement and density on the CNT surface and, consequently the availability of functional groups, varies considerably. An alternative approach to functionalize CNTs with an - additionally closed and precisely ordered - protein layer is offered by bacterial surface layer (S-layers) proteins which have already attracted much attention in the functionalization of surfaces as well as supporting structures for biomembranes. In a broad range of bacteria and archaea S-layer proteins cover the cells completely and may be considered as one of the most abundant biopolymers on earth. S-layer protein lattices show parameters in the nanometer range and offer surface chemical groups and genetically introduced biologically functional domains in precisely defined locations and orientation on their surfaces. Moreover, and highly relevant for this project too, is the natural capability of isolated S-layer proteins to self-assemble into monolayers in solution and at interfaces (e.g. on solid supports). The overall project aim is to conduct fundamental studies on the reassembly of S-layer proteins on CNTs and learn from nature how these new hybrid architectures may be used to make novel materials e.g. for biosensing. Key are the reassembly and binding properties of S-layer proteins which allow a highly specific and sensitive functionalization of the CNT surface. Moreover, novel hybrid organic-inorganic nano structures (e.g. nano containers for drug delivery) will become possible by using the S-layer coating as template in the biomineralization of silica, metals or other technologically important materials. Further on, it may also be assumed that the pores in the S-layer lattice will induce an ordered arrangement of metallic nanoparticles directly on the CNT surface and thus might lead to new electronic effects along the “one-dimensional” CNTs. Based on these few examples of an S-layer protein and CNTs construction kit, we would like to stress that our research, although longer term in nature, might lead to a new technology for the functionalization of carbon nanotube surfaces.