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Author(s): Youxing Jiang [1]; Alice Lee; Jiayun Chen; Vanessa Ruta; Martine Cadene; Brian T. Chait; Roderick MacKinnon (corresponding author) Many cells produce electrical impulses, known as action potentials, that propagate [...]

Author(s): Youxing Jiang; Alice Lee; Jiayun Chen; Martine Cadene; Brian T. Chait; Roderick MacKinnon (corresponding author) Potassium and other ion channels are allosteric proteins that switch between closed and opened [...]

Author(s): Youxing Jiang; Alice Lee; Jiayun Chen; Martine Cadene; Brian T. Chait; Roderick MacKinnon (corresponding author) Ion channels are central to a wide range of biological processes including cell volume [...]

Author(s): Raimund Dutzler [1]; Ernest B. Campbell [1]; Martine Cadene [2]; Brian T. Chait [2]; Roderick MacKinnon (corresponding author) [1] Potassium, sodium, calcium and chloride ions are used ingeniously by [...]

Research reveals that the endoplasm reticulum glycoprotein gp96/GRP94 oligomerizes into larger multimeric complexes with size ranging from dimers to octamers and higher. The oligomerization occurs with or without the dimerization domain participation, and in the peptide uptake by gp96, the dimerization domain plays only a minor role.

Research has been conducted on the covalent association of the latent TGF-beta binding protein-1 to latent TGF-beta. A recovery of functional recombinant GR3 is discussed.

Research was conducted to examine the inhibition of cathepsin G by native and oxidized mucus proteinase inhibitor (MPI) in the absence and presence of heparin. The stopped-flow kinetic investigation reveals that, within the inhibitor concentration range used, the mechanism of inhibition of cathepsin G an chymotrypsin by oxidized MPI is consistent with a one-step reaction while the inhibition of elastase takes place in two steps. Results demonstrate that oxidation of MPI during chronic bronchitis may result in cathepsin G-mediated lung tissue degradation.

The neutrophil elastase (NE)-suramin interaction was studied through steady-state and time-resolved fluorescence spectroscopy to the determine the suramin molecules on NE. Data from the longest lifetime class in the presence of variable suramin concentrations revealed that two suramins are bound at less than 17 angstroms from Trp 141 while the other two are found at least 29 angstroms from Trp 141. Also, suramin does not directly interfere with the substrate binding site of NE.

Heparin attaches to mucus proteinase inhibitor (MPI) in the reference aqueous medium through ionic reactions of weak energy with an enthalpy per site of about -10 kj per mole-1. The total enthalpy regulates the association process and, though the individual reactions are weak, the total enthaply is big. Heparin changes the inhibitor's activity through its agonist ligand behavior when attaching to the MPI receptor.

The total rate at which the enzyme and its inhibitor associate with one another falls due to the influence of heparin. This is due to the two-stage process through which the alpha-1-proteinase inhibitor (alpha 1 PI) reacts with the human neutrophil elastase (NE)-heparin complex with the employment of slow-binding kinetics. The interaction results in the emission of a pre-steady state release produce in association with a substrate. The enzyme is adsorbed to the spectro photometer, causing the steady-state velocity. The binding cannot be altered, as proved by the resistance of the alpha-1 PI-NE and the alpha 1a PI-NE heparin complex to dilution and dissociation.

Polymorphonuclear neutrophils contain at least four serine endopeptidases, namely neutrophil elastase (NE), proteinase 3 (PR3), cathepsin G (CatG), and NSP4, which contribute to the regulation of infection and of inflammatory processes. In physiological conditions, endogenous inhibitors including α2‐macroglobulin (α2‐M), serpins [α1‐proteinase inhibitor (α1‐PI)], monocyte neutrophil elastase inhibitor (MNEI), α1‐antichymotrypsin, and locally produced chelonianins (elafin, SLPI) control excessive proteolytic activity of neutrophilic serine proteinases. In contrast to human NE (hNE), hPR3 is weakly inhibited by α1‐PI and MNEI but not by SLPI. α2‐M is a large spectrum inhibitor that traps a variety of proteinases in response to cleavage(s) in its bait region. We report here that α2‐M was more rapidly processed by hNE than hPR3 or hCatG. This was confirmed by the observation that the association between α2‐M and hPR3 is governed by a kass in the ≤ 105 m−1·s−1 range. Since α2‐M‐trapped proteinases retain peptidase activity, we first predicted the putative cleavage sites within the α2‐M bait region (residues 690–728) using kinetic and molecular modeling approaches. We then identified by mass spectrum analysis the cleavage sites of hPR3 in a synthetic peptide spanning the 39‐residue bait region of α2‐M (39pep‐α2‐M). Since the 39pep‐α2‐M peptide and the corresponding bait area in the whole protein do not contain sequences with a high probability of specific cleavage by hPR3 and were indeed only slowly cleaved by hPR3, it can be concluded that α2‐M is a poor inhibitor of hPR3. The resistance of hPR3 to inhibition by endogenous inhibitors explains at least in part its role in tissue injury during chronic inflammatory diseases and its well‐recognized function of major target autoantigen in granulomatosis with polyangiitis. [ABSTRACT FROM AUTHOR]

Recent breakthroughs in the high-resolution structural elucidation of ion channels and transporters are prompting a growing interest in methods for characterizing integral membrane proteins. These methods are proving extremely valuable in facilitating the production of X-ray diffraction-grade crystals. Here we present a robust and straightforward mass spectrometric procedure that utilizes matrix-assisted laser desorption/ionization to analyze integral membrane proteins in the presence of detergents. The utility of this method is illustrated with examples of high-quality mass spectral data obtained from membrane proteins for which atomic resolution structural studies are ongoing.