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15. The conformations and basal conformational dynamics of translocation factor SecDF vary with translocon SecYEG interaction.

Author

Weaver, D. R., Amin, D. N., and King, G. M.

Subjects
*PROTEIN precursors, *MEMBRANE proteins, *ATOMIC force microscopy, *PROTEIN conformation, *BILAYER lipid membranes, *ESCHERICHIA coli
Abstract

The general secretory, or Sec, system is a primary protein export pathway from the cytosol of Escherichia coli and all eubacteria. Integral membrane protein complex SecDF is a translocation factor that enhances polypeptide secretion, which is driven by the Sec translocase, consisting of translocon SecYEG and ATPase SecA. SecDF is thought to utilize a proton gradient to effectively pull precursor proteins from the cytoplasm into the periplasm. Working models have been developed to describe the structure and function of SecDF, but important mechanistic questions remain unanswered. Atomic force microscopy (AFM) is a powerful technique for studying the dynamics of single-molecule systems including membrane proteins in near-native conditions. The sharp tip of the AFM provides direct access to membrane-external protein conformations. Here, we acquired AFM images and kymographs (~100 ms resolution) to visualize SecDF protrusions in near-native supported lipid bilayers and compared the experimental data to simulated AFM images based on static structures. When studied in isolation, SecDF exhibited a stable and compact conformation close to the lipid bilayer surface, indicative of a resting state. Interestingly, upon SecYEG introduction, we observed changes in both SecDF conformation and conformational dynamics. The population of periplasmic protrusions corresponding to an intermediate form of SecDF, which is thought to be active in precursor protein handling, increased more than ninefold. In conjunction, our dynamics measurements revealed an enhancement in the transition rate between distinct SecDF conformations when the translocon was present. Together, this work provides a novel vista of basal-level SecDF conformational dynamics in near-native conditions. [ABSTRACT FROM AUTHOR]

Published
2022
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23. Symposia

Author

Jordan, A., Eckert, D., Wellman, A., Malhotra, A., White, D., Saboisky, J., Lo, Y.-L., Moore, B., Bruck, D., Walsh, J., Maddison, K., Tesfayesus, W., Hillman, D., Eastwood, P., Scarfe, W., Wirtz-Justice, A., Lack, L., Burgess, H., Antic, N., Anderson, V., Kohler, M., Mcevoy, D., White, D. P., Horne, R., Hamilton, G., Weaver, D., Rajaratnam, S. M. W., Singh, B., Hill, N., Nixon, G., Howard, M., Harvey, A. S., Hiscock, H., Quach, J., Wake, M., Cain, N., Gradisar, M., Moseley, L., Paine, S., Proietto, J., Archer, J., Swann, P., Ferguson, S., Gander, P., Signal, L., Landrigan, C. P., Czeisler, C. A., Wilson, J., Hare, D., and Pierce, R.

Published
2009
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26. Influence of Protein Glycosylation onCampylobacter fetusPhysiology

Author

Duma, J, Nothaft, H, Weaver, D, Fodor, C, Beadle, B, Linton, D, Benoit, SL, Scott, NE, Maier, RJ, Szymanski, CM, Duma, J, Nothaft, H, Weaver, D, Fodor, C, Beadle, B, Linton, D, Benoit, SL, Scott, NE, Maier, RJ, and Szymanski, CM

Abstract

Campylobacter fetus is commonly associated with venereal disease and abortions in cattle and sheep, and can also cause intestinal or systemic infections in humans that are immunocompromised, elderly, or exposed to infected livestock. It is also believed that C. fetus infection can result from the consumption or handling of contaminated food products, but C. fetus is rarely detected in food since isolation methods are not suited for its detection and the physiology of the organism makes culturing difficult. In the related species, Campylobacter jejuni, the ability to colonize the host has been linked to N-linked protein glycosylation with quantitative proteomics demonstrating that glycosylation is interconnected with cell physiology. Using label-free quantitative (LFQ) proteomics, we found more than 100 proteins significantly altered in expression in two C. fetus subsp. fetus protein glycosylation (pgl) mutants (pglX and pglJ) compared to the wild-type. Significant increases in the expression of the (NiFe)-hydrogenase HynABC, catalyzing H2-oxidation for energy harvesting, correlated with significantly increased levels of cellular nickel, improved growth in H2 and increased hydrogenase activity, suggesting that N-glycosylation in C. fetus is involved in regulating the HynABC hydrogenase and nickel homeostasis. To further elucidate the function of the C. fetus pgl pathway and its enzymes, heterologous expression in Escherichia coli followed by mutational and functional analyses revealed that PglX and PglY are novel glycosyltransferases involved in extending the C. fetus hexasaccharide beyond the conserved core, while PglJ and PglA have similar activities to their homologs in C. jejuni. In addition, the pgl mutants displayed decreased motility and ethidium bromide efflux and showed an increased sensitivity to antibiotics. This work not only provides insight into the unique protein N-glycosylation pathway of C. fetus, but also expands our knowledge on the influence of pr

Published
2020

30. Control of VR-7 dynamic stall by strong steady blowing

Author

Weaver, D., McAlister, K.W., and Tso, J.

Subjects
Aerofoils -- Research, Aerospace and defense industries, Business, Science and technology
Abstract

An experiment was performed in a water tunnel on a Boeing-Vertol VR-7 airfoil to study the effects of tangential blowing over the upper surface. Blowing was applied at the quarter-chord location during sinusoidal pitching oscillations described by [alpha] = [[alpha].sub.m] + 10 deg sin [omega]t. Results were obtained for a Reynolds number of 1 x [10.sup.5], mean angles of 10 and 15 deg, reduced frequencies ranging from 0.005 to 0.15, and blowing rates from [C.sub.[micro]] =0.16 to 0.66. Unsteady lift, drag, and pitching moment loads are reported, along with fluorescent-dye flow visualizations. Strong steady blowing was found to prevent the bursting of the leading-edge separation bubble at several test points. When this occurred, the lift was increased significantly, stall was averted, and the shape of the moment response showed a positive damping in pitch. In almost all cases, steady blowing reduced the hysteresis amplitudes present in the loads, but the benefits diminished as the reduced frequency and mean angle of oscillation increased. A limited number of pulsed blowing cases indicated that for low blowing rates, the greatest gains were achieved at [F.sup.+] = 0.9.

Published
2004

33. Albedo and atmospheric constraints of dwarf planet Makemake from a stellar occultation

Author

Ortiz, J. L., Sicardy, B., Braga-Ribas, F., Alvarez-Candal, A., Lellouch, E., Duffard, R., Pinilla-Alonso, N., Ivanov, V. D., Littlefair, S. P., Camargo, J. I. B., Assafin, M., Unda-Sanzana, E., Jehin, E., Morales, N., Tancredi, G., Gil-Hutton, R., de la Cueva, I., Colque, J. P., Da Silva Neto, D. N., Manfroid, J., Thirouin, A., Gutiérrez, P. J., Lecacheux, J., Gillon, M., Maury, A., Colas, F., Licandro, J., Mueller, T., Jacques, C., Weaver, D., Milone, A., Salvo, R., Bruzzone, S., Organero, F., Behrend, R., Roland, S., Vieira-Martins, R., Widemann, T., Roques, F., Santos-Sanz, P., Hestroffer, D., Dhillon, V. S., Marsh, T. R., Harlingten, C., Bagatin, Campo A., Alonso, M. L., Ortiz, M., Colazo, C., Lima, H. J. F., Oliveira, A. S., Kerber, L. O., Smiljanic, R., Pimentel, E., Giacchini, B., Cacella, P., and Emilio, M.

Published
2012
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36. A Pluto-like radius and a high albedo for the dwarf planet Eris from an occultation

Author

Sicardy, B., Ortiz, J. L., Assafin, M., Jehin, E., Maury, A., Lellouch, E., Hutton, Gil R., Braga-Ribas, F., Colas, F., Hestroffer, D., Lecacheux, J., Roques, F., Santos-Sanz, P., Widemann, T., Morales, N., Duffard, R., Thirouin, A., Castro-Tirado, A. J., Jelínek, M., Kubánek, P., Sota, A., Sánchez-Ramírez, R., Andrei, A. H., Camargo, J. I. B., da Silva Neto, D. N., Gomes, Ramos A., Martins, Vieira R., Gillon, M., Manfroid, J., Tozzi, G. P., Harlingten, C., Saravia, S., Behrend, R., Mottola, S., Melendo, García E., Peris, V., Fabregat, J., Madiedo, J. M., Cuesta, L., Eibe, M. T., Ullán, A., Organero, F., Pastor, S., de los Reyes, J. A., Pedraz, S., Castro, A., de la Cueva, I., Muler, G., Steele, I. A., Cebrián, M., Montañés-Rodríguez, P., Oscoz, A., Weaver, D., Jacques, C., Corradi, W. J. B., Santos, F. P., Reis, W., Milone, A., Emilio, M., Gutiérrez, L., Vázquez, R., and Hernández-Toledo, H.

Published
2011
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37. Clinical evaluation of cardiovascular devices: principles, problems, and proposals for European regulatory reform: Report of a policy conference of the European Society of Cardiology†

Author

Fraser, Alan G., Daubert, Jean-Claude, Van de Werf, Frans, Estes, N.A. Mark, III, Smith, Sidney C., Jr, Krucoff, Mitchell W., Vardas, Panos E., Komajda, Michel, Anker, SD, Auricchio, A, Bailey, S, Bonhoeffer, P, Borggrefe, M, Brodin, L-Å, Bruining, N, Buser, P, Butchart, E, Calle Gordo, J, Cleland, J, Danchin, N, Daubert, JC, Degertekin, M, Demade, I, Denjoy, N, Derumeaux, G, Di Mario, C, Dickstein, K, Dudek, D, Estes, NA, Farb, A, Flotats, A, Fraser, AG, Gueret, P, Israel, C, James, S, Kautzner, J, Komajda, M, Krucoff, MW, Lombardi, M, Marwick, T, Mioulet, M, OʼKelly, S, Perrone-Filardi, P, Rosano, G, Rosenhek, R, Sabate, M, Smith, SC, Swahn, E, Tavazzi, L, Van de Werf, F, van der Velde, E, van Herwerden, L, Vardas, PE, Voigt, J-U, Weaver, D, and Wilmshurst, P.

Published
2011
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38. Methods for detecting PER2::LUCIFERASE bioluminescence rhythms in freely moving mice

Author

Martin-Burgos, B., primary, Wang, W., additional, William, I., additional, Tir, S., additional, Mohammad, I., additional, Javed, R., additional, Smith, S., additional, Cui, Y., additional, Smith, C.B., additional, van der Vinne, V., additional, Molyneux, P.C., additional, Miller, S.C., additional, Weaver, D. R., additional, Leise, T.L., additional, and Harrington, M.E., additional

Published
2020
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