ChemEM: Flexible Docking of Small Molecules in Cryo-EM Structures.

Cryo-electron microscopy (cryo-EM), through resolution advancements, has become pivotal in structure-based drug discovery. However, most cryo-EM structures are solved at 3-4 Å resolution, posing challenges for small-molecule docking and structure-based virtual screening due to issues in the precise positioning of ligands and the surrounding side chains. We present ChemEM, a software package that employs cryo-EM data for the accurate docking of one or multiple ligands in a protein-binding site. Validated against a highly curated benchmark of high- and medium-resolution cryo-EM structures and the corresponding high-resolution controls, ChemEM displayed impressive performance, accurately placing ligands in all but one case, often surpassing cryo-EM PDB-deposited solutions. Even without including the cryo-EM density, the ChemEM scoring function outperformed the well-established AutoDock Vina score. Using ChemEM, we illustrate that valuable information can be extracted from maps at medium resolution and underline the utility of cryo-EM structures for drug discovery.

Local regularization of tilt projections reduces artifacts in electron tomography.

Electron tomography produces very high resolution 3D image volumes useful for investigating the structure and function of cellular components. Unfortunately, unavoidable discontinuities and physical constraints in the acquisition geometry lead to a range of artifacts that can affect the reconstructed image. In particular, highly electron dense regions, such as gold nanoparticles, can hide proximal biological structures and degrade the overall quality of the reconstructed tomograms. In this work we introduce a pre-reconstruction non-conservative non-linear isotropic diffusion (NID) filter that automatically identifies and reduces local irregularities in the tilt projections. We illustrate the improvement in quality obtained using this approach for reconstructed tomograms generated from samples of malaria parasite-infected red blood cells. A quantitative and qualitative evaluation for our approach on both simulated and real data is provided.

Spatial and temporal mapping of the PfEMP1 export pathway in Plasmodium falciparum.

The human malaria parasite, Plasmodium falciparum, modifies the red blood cells (RBCs) that it infects by exporting proteins to the host cell. One key virulence protein, P. falciparum Erythrocyte Membrane Protein-1 (PfEMP1), is trafficked to the surface of the infected RBC, where it mediates adhesion to the vascular endothelium. We have investigated the organization and development of the exomembrane system that is used for PfEMP1 trafficking. Maurer's cleft cisternae are formed early after invasion and proteins are delivered to these (initially mobile) structures in a temporally staggered and spatially segregated manner. Membrane-Associated Histidine-Rich Protein-2 (MAHRP2)-containing tether-like structures are generated as early as 4 h post invasion and become attached to Maurer's clefts. The tether/Maurer's cleft complex docks onto the RBC membrane at ~20 h post invasion via a process that is not affected by cytochalasin D treatment. We have examined the trafficking of a GFP chimera of PfEMP1 expressed in transfected parasites. PfEMP1B-GFP accumulates near the parasite surface, within membranous structures exhibiting a defined ultrastructure, before being transferred to pre-formed mobile Maurer's clefts. Endogenous PfEMP1 and PfEMP1B-GFP are associated with Electron-Dense Vesicles that may be responsible for trafficking PfEMP1 from the Maurer's clefts to the RBC membrane.

Improving the quality of electron tomography image volumes using pre-reconstruction filtering.

Electron tomography produces highly magnified 3D image volumes useful for investigating the structure and function of cellular components. Image quality is degraded by multiple scattering events and quantum noise, which depend on the angle at which individual tilt projections are collected. We have adapted a biomedical imaging approach to improve image quality by enhancing individual tilt projections prior to volumetric reconstruction. Specifically, we have developed a family of non-linear anisotropic diffusion (NAD) filters parameterized by the tilt angle. We give a quantitative and qualitative evaluation of our pre-processing approach and the NAD filter. We show an improvement in the reconstructed volumes for tomograms generated from both plastic-embedded and cryo-stabilized samples of malaria parasite-infected erythrocytes.