The serum level of iron in humans is tightly controlled by the action of the hormone hepcidin on the iron efflux transporter ferroportin Hepcidin regulates iron absorption and recycling by inducing the internalization and degradation of ferroportin1 Aberrant ferroportin activity can lead to diseases of iron overload, such as haemochromatosis, or iron limitation anaemias2 Here we determine cryogenic electron microscopy structures of ferroportin in lipid nanodiscs, both in the apo state and in complex with hepcidin and the iron mimetic cobalt These structures and accompanying molecular dynamics simulations identify two metal-binding sites within the N and C domains of ferroportin Hepcidin binds ferroportin in an outward-open conformation and completely occludes the iron efflux pathway to inhibit transport The carboxy terminus of hepcidin directly contacts the divalent metal in the ferroportin C domain Hepcidin binding to ferroportin is coupled to iron binding, with an 80-fold increase in hepcidin affinity in the presence of iron These results suggest a model for hepcidin regulation of ferroportin, in which only ferroportin molecules loaded with iron are targeted for degradation More broadly, our structural and functional insights may enable more targeted manipulation of the hepcidin-ferroportin axis in disorders of iron homeostasis

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Structure of hepcidin-bound ferroportin reveals iron homeostatic mechanisms.

This review describes selected basics of water in biomolecular recognition. We focus on a qualitative understanding of the most important physical aspects, how these change in magnitude between bulk water and protein environment, and how the roles that water plays for proteins arise from them. These roles include mechanical support, thermal coupling, dielectric screening, mass and charge transport, and the competition with a ligand for the occupation of a binding site. The presence or absence of water has ramifications that range from the thermodynamic binding signature of a single ligand up to cellular survival. The large inhomogeneity in water density, polarity and mobility around a solute is hard to assess in experiment. This is a source of many difficulties in the solvation of protein models and computational studies that attempt to elucidate or predict ligand recognition. The influence of water in a protein binding site on the experimental enthalpic and entropic signature of ligand binding is still a point of much debate. The strong water-water interaction in enthalpic terms is counteracted by a water molecule's high mobility in entropic terms. The complete arrest of a water molecule's mobility sets a limit on the entropic contribution of a water displacement process, while the solvent environment sets limits on ligand reactivity.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/jmr.2810

Water in protein hydration and ligand recognition.

α-tubulin tail modifications regulate microtubule stability through selective effector recruitment, not changes in intrinsic polymer dynamics

The voltage-dependent motor protein Prestin (SLC26A5) is responsible for the electromotive behavior of outer hair cells and underlies the cochlear amplifier1. Knock out or impairment of Prestin causes severe hearing loss2–5. Despite Prestin’s key role in hearing, the mechanism by which mammalian Prestin senses voltage and transduces it into cellular-scale movements (electromotility) is poorly understood. Here, we determined the structure of dolphin Prestin in six distinct states using single particle cryo-electron microscopy. Our structural and functional data suggest that Prestin adopts a unique and complex set of states, tunable by the identity of bound anions (Cl- or SO42-). Salicylate, a drug that can cause reversible hearing loss, competes for the anion-binding site of Prestin, and inhibits its function by immobilizing Prestin in a novel conformation. Our data suggests that the bound anion together with its coordinating charged residues and helical dipole act as a dynamic voltage sensor. Analysis of all anion-dependent conformations reveals how structural rearrangements in the voltage sensor are coupled to conformational transitions at the protein-membrane interface, suggesting a novel mechanism of area expansion. Visualization of Prestin’s electromotility cycle distinguishes Prestin from closely related SLC26 anion transporters, highlighting the basis for evolutionary specialization of the mammalian cochlear amplifier at high resolution.

Prestin's conformational cycle underlies outer hair cell electromotility

Water is often found to mediate interactions between a ligand and a protein. It can play a significant role in orientating the ligand within a binding pocket and contribute to the free energy of binding. It would thus be extremely useful to be able to accurately predict the position and orientation of water molecules within a binding pocket. Recently, we developed the WaterDock protocol that was able to predict 97% of the water molecules in a test set. However, this approach generated false positives at a rate of over 20% in most cases and whilst this might be acceptable for some applications, in high throughput scenarios this is not desirable. Here we tackle this problem via the inclusion of knowledge regarding the solvation structure of ligand functional groups. We call this new protocol WaterDock2 and demonstrate that this protocol maintains a similar true positive rate to the original implementation but is capable of reducing the false-positive rate by over 50%. To improve the usability of the method, we have also developed a plugin for the popular graphics program PyMOL. The plugin also contains an implementation of the original WaterDock.

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Waterdock 2.0: Water placement prediction for Holo-structures with a pymol plugin.

A new method of hydrating protein structures, which we call Dowser++, is presented. The method is based on a semi-empirical modification of a popular program for protein hydration Dowser, and the usage of protocols AutoDock Vina, and WaterDock. The positions of water molecules predicted by Dowser++ were compared with experimental data for a set of 14 high-resolution crystal structures of oligopeptide-binding protein (OppA) containing a large number of resolved internal water molecules, as well as for the D- and K-channels of cytochrome c oxidase, and the recent data on PSII. Comparison is also made with the predictions of the original Dowser, and its improved version, Dowser+, described in our previous publication. We also present a model for quantitative estimation of the quality of water molecules placement made by a program, which includes an assumption of possible false negative data from the crystallographic analysis. The comparison of predictions made by Dowser++, Dowser and Dowser+ demonstrates significant improvement of predictive power of the new method. Proteins 2016; 84:1347-1357. © 2016 Wiley Periodicals, Inc.

A. A. Stuchebrukhov

Papers

Dowser++, a new method of hydrating protein structures