We acknowledge the following funding:

SFB 1027

“Physical modeling of non-equilibrium processes in biological systems”

Gene expression in all biological cells is tightly regulated by the binding of transcription factors and by epigenetic modifications of the DNA. Importantly, gene expression and cell differentiation or cell reprogramming are triggered by suitable stimuli in a stochastic manner. Here, atomistic biomolecular simulations and coarse-grained Brownian dynamics simulations will be used to study the binding processes governing gene expression in the E.coli pap operon that is being studied experimentally in project C1. A second part of the project involves stochastic dynamics simulations to model state transitions of the gene-regulatory network centered on the pluripotency factors Oct4, Nanog and Sox2. In collaboration with project C2, we will characterize how dynamic changes of transcription factor concentrations and DNA methylation levels affect cell differentiation during the development of the early mouse embryo until the 32-cell stage.

More at http://

DFG Project

“How allosteric effectors influence protein translocation mediated by the Sec61 complex”

In eukaryotes, the protein biosynthesis is carried out either by cytosolic ribosomes or by ribosomes that are attached to the membrane of the endoplasmatic reticulum (ER). In the latter case, the newly synthesized “nascent peptide chain” (NC) is typically translocated into the ER or laterally inserted into the ER membrane via an integral membrane protein complex. In eukaryotes, its central component is termed the Sec61 complex. Proteins that should be translocated across the ER membrane carry a “signal Peptide” (SP) at their N-terminus. In the ER, this sequence is then cleaved off by the enzyme signal peptidase. Upon exiting from the ribosome, the signal recognition particle (SRP) binds SP of the newly synthesized NC at the ribosome. Subsequently, upon interaction of SRP and SRP receptor at the ER membrane, SRP dissociates from SP which then is free to insert into the Sec61 channel. This pathway is termed the co-translational, SRP-dependent pathway, which is the focus of the proposed work. Not all NCs that should pass Sec61 may do this on their own. Certain NCs require the presence of additional accessory membrane proteins such as TRAP and Sec62-Sec63.

Until recently, it was unclear which features of SPs guide them through different translocation pathways. In prior published work, we have contributed to unraveling physico chemical features of SPs that distinguish TRAP clients or Sec62-63 clients from those proteins that do not require aid by such accessory proteins. Still, the mechanistic role of these accessory proteins and how they mechanistically affect cotranslational translocation is unknown. Furthermore, several small molecule effectors have been described in the literature (e.g. eeyarestatins, mycolactone, and Ipomoeassin F) that modulate the protein translocation efficiency of the Sec61 complex. In prior published work, we combined ligand docking with data from electrophysiology to suggest how eeyarestatins derivatives bind differentially in the Sec61 pore. Based on our published findings and on recent advances in structure determination of the Sec61 complex, we now aim at unravelling the mechanistic details how allosteric effectors influence protein translocation mediated by the Sec61 complex. We will implement an integrated computational and experimental approach.

Specific aims of the proposed project are:

  1. We want to characterize how SPs bind in the Sec61 pore,
  2. we want to characterize how accessory proteins alter the conformational sampling of SP in the Sec61 pore,
  3. we want to characterize the molecular consequences upon binding of small-molecule effectors to Sec61.

DFG Project

“How phosphorylation affects peptide interaction with adaptor domains”

Protein-protein interactions (PPIs) are of fundamental relevance for most processes in biological cells and hence the focus of considerable experimental and computational efforts. Notably, PPIs may also be strongly affected by post-translational modifications of the involved proteins such as phosphorylation that may either favor or disfavor formation of a particular complex. In this project, we will analyze interactions of adaptor domains with phosphorylated and unphosphorylated peptide binders by means of molecular dynamics simulations. We will build on the successful characterization of such binding processes in a first funding phase, where we studied binding of phospho/non-phospho peptides to the PDZ2 domain of the hPTP1E protein and to the PDZ1 domain of the MAGI1 protein by comparable simulations. In this project, we will at first correct slight inaccuracies of the phosphate parameters by calibration against experimental binding affinities. In collaboration with a structural biology group, we aim at establishing a mechanistic description why phosphorylated amino acids can either favor or disfavor binding of the peptide to PDZ domains depending on where the phosphate group is placed along the peptide sequence. These MD simulations will then be extended to MAP1LC3A ATG8 domains and TSG101 UEV domains. Finally, we will compile a comprehensive dataset on complexes of adaptor domains with phosphorylated peptides. Based on this, we will develop classification systems based on machine learning that predict for a given atomistic structure of an adaptor domain and for the sequence of a peptide binding to it whether ist phosphorylated or nonphosphorylated variant is predicted to bind stronger and whether glutamic acid is a suitable phosphomimetic of phospho serine for this domain-peptide pair. We will start with developing classifiers for PDZ domains, followed by versions for general adaptor domains.