Research Interests

Gene regulation, DNA repair, structure and dynamics of proteins and complexes

Our research focuses on understanding the mechanism of gene regulation, expression and repair. Using NMR spectroscopy we study the structure and dynamics of protein and protein-DNA complexes involved in transcription, translation and DNA repair. Studied examples are the E. coli Lac repressor, bacterial ribosomal initiation factors, the human ERCC1/XPF DNA repair complex and the mechanism of chromatin remodeling. Further studied systems include the photoreceptors PYP and AppA, the Kid/Kis plasmid maintenance system and enzymes, such as subtilisins and lipases.

Specific and non-specific DNA binding by Lac repressor

The lac system is used as a model system to probe in detail structural and dynamic aspects of protein-DNA recognition. Our studies of the complex of Lac headpiece (the DNA binding domain) with different lac operators show the plasticity that both proteins and DNA exhibit in order to optimize interactions. Interaction of DNA-binding proteins with their operators can be preceded by binding to nonspecific DNA. This speeds up the search for the target site by several orders of magnitude. As a model for one-dimensional sliding we determined the solution structure and dynamics of the complex of Lac headpiece with nonspecific DNA.

Kalodimos et al, EMBO J. 21 (2002) 2866-76, Science 305 (2004) 386-9, Chem.Rev. 104 (2004) 3567-86; Romanuka et al, J Mol Biol 390 (2009) 478-8; Loth et al, J BNMR 56 (2013) 41-9

Nucleotide Excision DNA repair

In collaboration with Jan Hoeijmakers (Erasmus MC, Rotterdam) we study the heterodimeric complex of the C-terminal parts of the human Nucleotide Excision DNA repair proteins XPF and ERCC1. The structure explains how ERCC1 targets the endonuclease XPF to the DNA damage site. Subsequent studies of the central domain of ERCC1 accounts for binding of ERCC1/XPF at a ds/ssDNA junction near the DNA lesion.

Tripsianes et al, Structure 13 (2005) 1849-58; Das et al, Proteins 70 (2008) 1551–63, Structure 20 (2012) 667-75

Translation initiation

Bacteria require three initiation factors, IF1, IF2 and IF3, to start protein synthesis. In collaboration with Claudio Gualerzi (Camerino, Italy) we study the structure, function and dynamics of these initiation factors, the changes therein upon interaction with ribosomal subunits, and in GTP and GDP bound states.

Meunier et al, EMBO J 19 (2000) 1918-26; Guennegues et al, EMBO J 19 (2000) 5233-40; Milon et al, PNAS 103 (2006), 13962-7; Wienk et al, JBC 287 (2012), 10922-32

Biomolecular docking

In collaboration with Alexandre Bonvin (Utrecht University) we developed the program HADDOCK for modeling protein-protein complexes that makes use of biochemical and/or biophysical interaction data such as resulting from NMR titration experiments or mutagenesis data. We used it for modeling E2-E3 ubiquitination complexes such as the complex of the human transcription factor CNOT4 and the ubiquitin conjugating enzyme UbcH5B (collaboration with Marc Timmers, UMCU) and the ubiquitination complexes of Rad8 and Rad16 (collaboration with Titia Sixma, NKI, Amsterdam).

Dominguez et al., J Am Chem Soc 125(2003) 1731-37, Structure 12(2004) 633-44; Hibbert et al, PNAS 108 (2011) 5590-5; Huang et al, J Mol Biol 385 (2011), 507-19