Effects of Distal Mutations on the Structure, Dynamics and Catalysis of Human Monoacylglycerol Lipase

An understanding of how conformational dynamics modulates function and catalysis of human monoacylglycerol lipase (hMGL), an important pharmaceutical target, can facilitate the development of novel ligands with potential therapeutic value. Here, we report the discovery and characterization of an allosteric, regulatory hMGL site comprised of residues Trp-289 and Leu-232 that reside over 18 Å away from the catalytic triad. These residues were identified as critical mediators of long-range communication and as important contributors to the integrity of the hMGL structure. Nonconservative replacements of Trp-289 or Leu-232 triggered concerted motions of structurally distinct regions with a significant conformational shift toward inactive states and dramatic loss in catalytic efficiency of the enzyme. Using a multimethod approach, we show that the dynamically relevant Trp-289 and Leu-232 residues serve as communication hubs within an allosteric protein network that controls signal propagation to the active site, and thus, regulates active-inactive interconversion of hMGL. Our findings provide new insights into the mechanism of allosteric regulation of lipase activity, in general, and may provide alternative drug design possibilities

Classic Isomeric 1, 2-and 2, 1-nitrosonaphthols are Oximes in Solid State and Solutions

A detailed characterization by UV-visible, IR and NMR (1H, 13C, 15N) spectroscopies of two isomeric 1,2- and 2,1-nitrosonaphthols (1 and 2) has been carried out. The crystal structures of these iconic compounds, which were widely used in analytical chemistry in the past, were determined and evidenced their quinone-oxime nature in solid state. Compound 1 exists in the crystal as a syn- diastereomer in the closed form due to a strong intramolecular H-bond, while compound 2 is an anti- diastereomer forming a dimer with two intermolecular H-bonds. The oxime character of 1 and 2 in solutions was confirmed after careful studies of the NMR spectra of both compounds in solvents of different polarity and donor properties: CD2Cl2, acetone-d6 and DMSO-d6. An equilibrium state between oxime syn- and anti- isomers exists and can be modulated in a controlled way by changing media\u27s polarity. Both compounds were also prepared labeled with 15N (50%) for recording their NMR nitrogen-15 spectra, and for the identification/assignment of vibrations with the participation of the \u3eC=N-OH fragment. During synthesis of compound 1 a persistent, very polar, red-colored impurity 3 was detected, isolated and crystallographically characterized. It was found to be a rather unusual π-complex between the final compound 1 and its precursor 2-naphthol. The red color originates from the CT-band in the visible region of the UV/Vis-spectrum where the naphthol acts as a donor with 1 being an acceptor. Both are separated at 3.03 A

Preparation, Properties and Crystal Structure of \u3cem\u3eSyn\u3c/em\u3e-Isomer of 2,6-Dichlorophenyl-Cyanoxime, H(2,6-diCl-PhCO): Potent \u3cem\u3eCarbonyl Reductase\u3c/em\u3e Inhibitor

The oximino(2,6-dichlorophenyl)acetonitrile, H(2,6-diCl-PhCO) has been synthesized in a reasonably high yield of 60%, and characterized using a variety of physical, electrochemical, spectroscopic methods and X-ray analysis. This compound belongs to the family of cyanoximes; a new subclass of oximes with the general formula NC–C(=N–OH)–R (where R is an electron-withdrawing group) which recently emerged as new biologically active compounds. This cyanoxime represents a disubstituted arylcyanoxime that was found to be a powerful inhibitor of the Carbonyl Reductase enzyme involved in the developing of resistance to anticancer treatment, and the making of cardiotoxic derivatives of anthracyclines that are currently used in medicine. The oximino(2,6-dichlorophenyl)acetonitrile, H(2,6-diCl-PhCO) is a weak acid with pKa = 6.17 and does not dissociate in organic polar protic and aprotic solvents. The cyanoxime was obtained as a microcrystalline mixture of two diastereomers (anti- and syn-) and deprotonates in solutions with the formation of yellow anions which exhibit solvatochromic behavior. However, one specific diastereomer—syn—was isolated in crystalline form from a solvent system as colorless blocks overlayed with pentane ether solution in a monoclinic system in a P2/c (#13) space group with unit cell parameters:  = 8.1720(2),  = 8.8013(3),  = 13.0146(4) and β = 102.546(3); Z = 4. A single crystal was studied using filtered CuKa radiation, providing Rint value of 0.0348 from a full-sphere of reflections. A crystal structure was solved using direct methods, and well refined to R1 = 0.0459, wR2 = 0.1268 and GOF = 1.107. The studied specimen of oximino(2,6-dichlorophenyl)acetonitrile, H(2,6-diCl-PhCO), represents a highly non-planar, rare syn-diastereomer in which the oxime fragment is positioned towards the chlorinated phenyl group. In the crystal, the compound forms a columnar structure extended along the c-direction by using slipped π–π stacking interactions. Columns are interconnected via H-bonding between the oxime OH-group and N atom of the nitrile group with the following parameters: N–H = 1.841 Å, and 169.20° N···H–O angle. No thermal interconversion of syn- into anti- diastereomer was observed upon heating of crystals of one isomer under flow of Ar

Characterization of the neuron-specific L1-CAM cytoplasmic tail: naturally disordered in solution it exercises different binding modes for different adaptor proteins

L1, a highly conserved transmembrane glycoprotein member of the immunoglobulin superfamily of cell adhesion molecules, mediates many developmental processes in the nervous system. Here we present the biophysical characterization and the binding properties of the least structurally defined part of this receptor: its cytoplasmic tail (CT). We have shown by analytical ultracentrifugation and dynamic light scattering experiments that it is mostly monomeric and unstructured in aqueous solution. We have defined by nuclear magnetic resonance the molecular details of L1-CT binding to two major targets: a membrane-cytoskeletal linker (MCL), ezrin, and an endocytosis mediator, AP2. Surprisingly, in addition to the two previously identified ezrin binding motifs, the juxtamembrane and the (1176)YRSLE regions, we have discovered a third one, a part of which has been previously associated with binding to another MCL, ankyrin. For the L1 interaction with AP2 we have determined the precise interaction region surrounding the (1176)YRSLE binding site and that this overlaps with the second ezrin binding site. In addition, we have shown that the juxtamembrane region of L1-CT has some binding affinity to AP2-mu2, although the specificity of this interaction needs further investigation. These data indicate that L1-CT belongs to the class of intrinsically disordered proteins. Endogenous flexibility of L1-CT might play an important role in dynamic regulation of intracellular signaling: the ability of cytoplasmic tails to accommodate different targets has the potential to fine-tune signal transduction via cell surface receptors

Understanding hydrogen electrocatalysis by probing the hydrogen-bond network of water at the electrified Pt/solution interface

A grand challenge in electrochemistry is to understand and promote electrochemical processes by exploring and exploiting the interface. Herein, we promoted the hydrogen evolution and oxidation reactions (HER/HOR) of platinum (Pt) in base by introducing N-methylimidazoles into the Pt-water interface. In situ spectroscopic characterization of the interface together with Quantum Mechanics computations showed that this promotion is caused by the N-methylimidazoles facilitating diffusion of hydroxides across the interface by holding the second layer water close to Pt surfaces. We accordingly propose that the HER/HOR kinetics of Pt in acid and base is governed by diffusion of protons and hydroxides, respectively, through the hydrogen-bond network of interfacial water by the Grotthuss mechanism, which accounts for the pH-dependent HER/HOR kinetics of platinum, a long-standing puzzle. Moreover, we demonstrated a 40% performance improvement of an anion exchange membrane electrolyzer by adding 1,2-dimethylimidazole into the alkaline solution fed into its platinum cathode