As organisms become increasingly complex, so too must they evolve a more sophisticated molecular alphabet. The recent discovery of proteins that can adopt multiple structural states is one way of addressing this complexity and it has dramatically changed our view of the protein structure-function paradigm.

Almost 40% of the human proteome is predicted to consist of such proteins that contain long intrinsically disordered regions (IDRs) and therefore lack a stable, well-defined three-dimensional structure. Proteins with long IDRs are prevalent in cellular regulation and signalling processes, and are implicated in a vast array of diseases and pathologies.

By studying these proteins we aim to obtain a quantitative description of their dynamic structural ensembles to understand function. 

Transcription factors are particularly enriched in structural disorder where almost all of the ~1600 human factors contain long IDRs. Transcription factors usually have small and folded domains responsible for binding specific DNA sequences. The much longer IDRs contain the transcriptional activation domains and other regions important for binding other proteins and integrating the transcriptional machinery. Deciphering the sequence grammar of transcription factor IDRs and how it dictates structure, dynamics, and function, is a major goal in our lab.

A primary focus is on pioneer transcription factors; these can bind and open condensed chromatin, and initiate cell identity changes. We use single-molecule approaches to map the structure and dynamics of the factors to understand their interactions with chromatin, and we monitor their effects on chromatin structure. We then use biochemistry and synthetic biology to modulate their function. Finally, we observe their functions and reprogramming abilities using methods from in vitro to living cells. 

To study transcriptional regulation, we design and develop biochemical approaches for producing chromatin constructs, especially for single-molecule studies.

We are currently developing methods to site-specifically label nucleosome arrays with defined transcription factor binding sites, to monitor chromatin structure during transcription factor binding. Most chromatin research has relied on using very stable DNA sequences that are normally not found in organisms. 

We are therefore also devoted to developing approaches to use „native“ DNA sequences for nucleosome formation, to understand transcription under increasingly native-like conditions. 

Allostery is a biological mechanism fundamental for cells to maintain metabolic homeostasis by providing fast, fine-tuned control of enzyme-catalyzed metabolic reactions. In recent years, allosteric modulation of enzyme activities by small molecules has received a growing interest in the field of cancer metabolism, owing to various therapeutic benefits:

(i) Allosteric sites expand the range of potential targets, including enzymes that were previously thought undruggable.
(ii) High target-selectivity can be achieved as allosteric sites are weakly conserved and unique across homologous enzymes.
(iii) Allosteric modulators pose lower risk of toxicity because they do not compete with endogenous substrates.

Alteration in metabolism is often associated with diseases such as cancer. Among many metabolic alterations, enhanced glycolysis is the most prominent feature of various cancer types and offers a wealth of targets that are amenable to allosteric regulation.

However, not all enzymes in the glycolytic pathway are currently known to be allosterically modulated. Here, we intend to identify novel allosteric regulation in two glycolytic enzymes that are still termed non-allosteric: glucose-6-phosphate isomerase and triosephosphate isomerase. To this end, a biophysical screening strategy has been proposed for identification of hits from small-molecule libraries. The potential hits will be validated by functional assays and their allosteric binding sites will be elucidated using X-ray protein crystallography. Furthermore, the novel allosteric ligands will be optimized for higher binding affinity and selectivity by 3D-QSAR analysis.

The goal of this project is to investigate the mechanism of peptide inhibition on the epidermal growth factor receptor (EGFR) against cancer. Specific peptides against EGFR have been developed, and this receptor plays a significant role in cell proliferation in various cancer types including breast cancer. The binding sites of these peptides are not known, nor have the effects on the tyrosine kinase (TK) domain, which activates the receptor's signaling pathways, been explored. Additionally, little is known about the role of the intrinsically disordered C-terminus in the inhibition.

In this project, one of the objective is to elucidate the X-ray crystal structure of EGFR bound to the peptides to determine the precise binding site. Moreover, single-molecule FRET (Förster resonance energy transfer) methods will be applied to understand the functions of individual domains in receptor inhibition. Such an understanding of the action of these peptides opens the door to further drug development using in silico optimization methods, with their premise relying on the mapping of binding regions and the flexibility of the receptor.