Our research

Molecular machines

Many vital processes in the cell are catalysed by large macromolecular assemblies. These assemblies have been called molecular machines because just like machines from daily life they employ coordinated movements of separate parts to fulfill their often complicated tasks. The understanding of how molecular machines work is a strategic goal in modern molecular biology, but studying them is often challenging.

Fragile inter-molecular interactions make it typically difficult to purify intact molecular machines, while different functional states may be hard to separate biochemically. Consequently, purified samples of molecular machines often suffer from various extents of non-stoichiometric complex formation and/or conformational variability. The occurrence of multiple different structures, also called structural heterogeneity, poses problems for many tools in structural biology. It often interferes with crystallisation and decrease the effectiveness of biophysical techniques that study assemblies in bulk solution.

Three-dimensional electron microscopy (3D-EM) allows visualising single copies of molecular machines. Rapidly frozen in a thin layer of ice, these assemblies re free to adopt any of their functional states. Provided that images of distinct 3D structures can be separated in the computer, 3D-EM poses less stringent requirements on sample homogeneity than alternative techniques. In principle, from a single 3D-EM sample one may therefore obtain structural information about a range of "snapshots" along the functional cycle of these machines. Combining multiple snapshots into a movie of a functioning machine is then likely to improve our insights into its mechanism.

Amyloid filaments

In some diseases, protein molecules that are the building blocks of molecular machines misfold and end up in filamentous aggregates called amyloids. Amyloid filaments are observed in many neurodegenerative diseases, with Alzheimer's disease being the most common one. In Alzheimer's disease, two proteins aggregate to form amyloid filaments: amyloid-beta peptides form extracellular plaques, and tau forms intracellular tangles. Besides studying protein structures in their 'healthy' state inside molecular machines, we are also interested in their structure inside amyloid filaments.

Our approach

We develop methods to visualise intact molecular machines as well as amyloid filaments by 3D-EM. Previous work has mainly focussed on novel image processing techniques, in particular maximum likelihood and empirical Bayesian algorithms. Current investigations also comprise developments in sample preparation and data collection schemes.

We introduced various statistical image processing algorithms to the field of 3D-EM. We showed that a 3D maximum-likelihood classification approach can separate projection images of distinct conformations without the need for a priori knowledge about the structural heterogeneity in the data. We also introduced a novel, Bayesian view on the single-particle reconstruction problem that provides a convenient statistical framework, in which many aspects that were previously regarded as separate steps all come together in optimising a single, regularised likelihood function. An important advantage of this approach is that most of the corresponding parameters are estimated from the data themselves, so that user expertise is no longer required in obtaining cutting-edge reconstructions. We also extended this Bayesian framework to deal with the helical symmetry that is present in amyloid filaments.

Our methods developments are often driven by challenging structure determination projects in our own group or in collaboration with others. Through these projects, we have been able to reach near-atomic resolution reconstructions for a variety of different samples. Two example applications related to Alzheimer's disease are the human gamma-secretase complex, which cuts amyloid-precursor protein to form amyloid-beta peptides, and the tau filaments from the brain of an individual with Alzheimer's disease. Please see our structures and publications for many more applications.