Transport across membranes via electrostatic 'charge zippers'

• Applicants: Professor Dr. Marcus Elstner; Professorin Dr. Anne Ulrich
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms ; Bioinformatics and Theoretical Biology ; Biophysics ; Structural Biology ; Theoretical Chemistry: Molecules, Materials, Surfaces
• Term: from 2013 to 2018
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 246585620

We have recently identified 'charge zippers' as a novel structural motif in membrane proteins [Walther et al., Cell, 2013]. In such systems the charged amino acids are aligned in a quasi-symmetrical, complementary pattern along the protein sequence. This suggests that long ladders of salt bridges can be formed between amphiphilic alpha-helical segments. By spanning the hydrophobic lipid bilayer, these charge zippers could selectively transport protons across membranes or lead to the assembly of larger ion-conducting pores. The 3D structures and oligomeric architectures of two peptide systems, for which we have predicted such motif, shall be characterized here, in order to analyze the functional relevance of the postulated charge zippers. First, we focus on the bacterial peptide TisB (29 amino acids), which is produced in E. coli under stress and induces the formation of biofilms. Our preliminary functional assays and solid-state NMR analysis have shown that the insertion of TisB into a membrane leads to a controlled depletion of the proton gradient. First coarse-grained and all-atom simulations support our model, according to which TisB assembles into antiparallel dimers via 4 salt bridges and is thus able to selectively transport protons (oder OH- ions) across the membrane. As an experimental proof of concept we plan to use solid-state NMR distance measurements and spin-counting of the monomers to demonstrate that and study in detail how the expected salt bridges are formed. In parallel, molecular dynamics simulations with empirical force fields and calculations in statistical thermodynamics will be employed. Based on the free energy profiles of the ions during transport via a TisB dimer it should be possible to discriminate the different possible mechanisms. Combined quantum mechanical/molecular mechanical methods shall yield a detailed description of the local interactions.In the second part, we will employ the experimental and computational methods that have been established with TisB further to study the antimicrobial peptide Dermicdin from human sweat. With a length of 48 amino acids, this amphiphilic helical sequence binds initially to the surface of bacterial membranes, as we have shown using oriented circular dichroism. According to the charge zipper model, up to 7 intra- and intermolecular salt bridges could then be formed if the protein folds as a helical hairpin and assembles into an oligomeric pore. Based on sterical arguments, Dermcidin would then possess the optimal length to span a lipid bilayer and kill the bacteria by high ion conductivity.

• DFG Programme: Research Grants