Inhibitor-Induced Dimerization of an Essential Oxidoreductase from African Trypanosomes
Trypanosomal and leishmanial infections claim tens of thousands of lives each year. The metabolism of these unicellular eukaryotic parasites differs from the human host and their enzymes thus constitute promising drug targets. Tryparedoxin (Tpx) from Trypanosoma brucei is the essential oxidoreductase in the parasite's hydroperoxide-clearance cascade. In vitro and in vivo functional assays show that a small, selective inhibitor efficiently inhibits Tpx. With X-ray crystallography, SAXS, analytical SEC, SEC-MALS, MD simulations, ITC, and NMR spectroscopy, we show how covalent binding of this monofunctional inhibitor leads to Tpx dimerization. Intra- and intermolecular inhibitor-inhibitor, pro…
Predicting 19F NMR Chemical Shifts: A Combined Computational and Experimental Study of a Trypanosomal Oxidoreductase–Inhibitor Complex
Abstract The absence of fluorine from most biomolecules renders it an excellent probe for NMR spectroscopy to monitor inhibitor–protein interactions. However, predicting the binding mode of a fluorinated ligand from a chemical shift (or vice versa) has been challenging due to the high electron density of the fluorine atom. Nonetheless, reliable 19F chemical‐shift predictions to deduce ligand‐binding modes hold great potential for in silico drug design. Herein, we present a systematic QM/MM study to predict the 19F NMR chemical shifts of a covalently bound fluorinated inhibitor to the essential oxidoreductase tryparedoxin (Tpx) from African trypanosomes, the causative agent of African sleepi…
Inhibitor-induzierte Dimerisierung einer essentiellen Oxidoreduktase aus afrikanischen Trypanosomen
Structure of the Human TRPML2 Ion Channel Extracytosolic/Lumenal Domain.
Summary TRPML2 is the least structurally characterized mammalian transient receptor potential mucolipin ion channel. The TRPML family hallmark is a large extracytosolic/lumenal domain (ELD) between transmembrane helices S1 and S2. We present crystal structures of the tetrameric human TRPML2 ELD at pH 6.5 (2.0 A) and 4.5 (2.95 A), corresponding to the pH values in recycling endosomes and lysosomes. Isothermal titration calorimetry shows Ca2+ binding to the highly acidic central pre-pore loop which is abrogated at low pH, in line with a pH-dependent channel regulation model. Small angle X-ray scattering confirms the ELD dimensions in solution. Changes in pH or Ca2+ concentration do not affect…
CCDC 1862408: Experimental Crystal Structure Determination
Related Article: Annika Wagner, Thien Anh Le, Martha Brennich, Philipp Klein, Nicole Bader, Erika Diehl, Daniel Paszek, A. Katharina Weickhmann, Natalie Dirdjaja, R. Luise Krauth-Siegel, Bernd Engels, Till Opatz, Hermann Schindelin, Ute A. Hellmich|2019|Angew.Chem.,Int.Ed.|58|3640|doi:10.1002/anie.201810470