Search

Your search keyword '"Zschörnig Olaf"' showing total 124 results
Start Over Author "Zschörnig Olaf"
124 results on '"Zschörnig Olaf"'

1. Characterization of cationic lipid DNA transfection complexes differing in susceptability to serum inhibition

Author

Überla Klaus, Nchinda Godwin, and Zschörnig Olaf

Subjects
Biotechnology, TP248.13-248.65
Abstract

Abstract Background Cationic lipid DNA complexes based on DOTAP (1,2-dioleoyl-3-(trimethyammonium) propane) and mixtures of DOTAP and cholesterol (DC) have been previously optimized for transfection efficiency in the absence of serum and used as a non-viral gene delivery system. To determine whether DOTAP and DC lipid DNA complexes could be obtained with increased transfection effciency in the presence of high serum concentrations, the composition of the complexes was varied systematically and a total of 162 different complexes were analyzed for transfection efficiency in the presence and absence of high serum concentrations. Results Increasing the ratio of DOTAP or DC to DNA led to a dose dependent enhancement of transfection efficiency in the presence of high serum concentrations up to a ratio of approximately 128 nmol lipid/μg DNA. Transfection efficiency could be further increased for all ratios of DOTAP and DC to DNA by addition of the DNA condensing agent protamine sulfate (PS). For DOTAP DNA complexes with ratios of ≤ 32 nmol/μg DNA, peak transfection efficiencies were obtained with 4 μg PS/μg DNA. In contrast, increasing the amount of PS of DC complexes above 0.5 μg PS /μg DNA did not lead to significant further increases in transfection efficiency in the presence of high serum concentrations. Four complexes, which had a similar high transfection efficiency in cell culture in the presence of low serum concentrations but which differed largely in the lipid to DNA ratio and the amount of PS were selected for further analysis. Intravenous injection of the selected complexes led to 22-fold differences in transduction efficiency, which correlated with transfection efficiency in the presence of high serum concentrations. The complex with the highest transfection efficiency in vivo consisted of 64 nmol DC/ 16 μg PS/ μg DNA. Physical analysis revealed a predicted size of 440 nm and the highest zeta potential of the complexes analyzed. Conclusions Optimization of cationic lipid DNA complexes for transfection efficiency in the presence of high concentrations of serum led to the identification of a DC complex with high transduction efficiency in mice. This complex differs from previously described ones by higher lipid to DNA and PS to DNA ratios. The stability of this complex in the presence of high concentrations of serum and its high transduction efficiency in mice suggests that it is a promising candidate vehicle for in vivo gene delivery.

Published
2002
Full Text
View/download PDF

18. Characterization of cationic lipid DNA transfection complexes differing in susceptability to serum inhibition

Author

Nchinda, Godwin, Überla, Klaus, and Zschörnig, Olaf

Subjects
Serum, Recombinant Fusion Proteins, lcsh:Biotechnology, Transfection, Cell Line, Fatty Acids, Monounsaturated, Mice, Cations, lcsh:TP248.13-248.65, Animals, Humans, Luciferases, Lung, Mice, Inbred BALB C, Dose-Response Relationship, Drug, DNA, Lipids, Culture Media, Quaternary Ammonium Compounds, Cholesterol, Injections, Intravenous, Liposomes, Cattle, Female, Research Article, Plasmids
Abstract

Background Cationic lipid DNA complexes based on DOTAP (1,2-dioleoyl-3-(trimethyammonium) propane) and mixtures of DOTAP and cholesterol (DC) have been previously optimized for transfection efficiency in the absence of serum and used as a non-viral gene delivery system. To determine whether DOTAP and DC lipid DNA complexes could be obtained with increased transfection effciency in the presence of high serum concentrations, the composition of the complexes was varied systematically and a total of 162 different complexes were analyzed for transfection efficiency in the presence and absence of high serum concentrations. Results Increasing the ratio of DOTAP or DC to DNA led to a dose dependent enhancement of transfection efficiency in the presence of high serum concentrations up to a ratio of approximately 128 nmol lipid/μg DNA. Transfection efficiency could be further increased for all ratios of DOTAP and DC to DNA by addition of the DNA condensing agent protamine sulfate (PS). For DOTAP DNA complexes with ratios of ≤ 32 nmol/μg DNA, peak transfection efficiencies were obtained with 4 μg PS/μg DNA. In contrast, increasing the amount of PS of DC complexes above 0.5 μg PS /μg DNA did not lead to significant further increases in transfection efficiency in the presence of high serum concentrations. Four complexes, which had a similar high transfection efficiency in cell culture in the presence of low serum concentrations but which differed largely in the lipid to DNA ratio and the amount of PS were selected for further analysis. Intravenous injection of the selected complexes led to 22-fold differences in transduction efficiency, which correlated with transfection efficiency in the presence of high serum concentrations. The complex with the highest transfection efficiency in vivo consisted of 64 nmol DC/ 16 μg PS/ μg DNA. Physical analysis revealed a predicted size of 440 nm and the highest zeta potential of the complexes analyzed. Conclusions Optimization of cationic lipid DNA complexes for transfection efficiency in the presence of high concentrations of serum led to the identification of a DC complex with high transduction efficiency in mice. This complex differs from previously described ones by higher lipid to DNA and PS to DNA ratios. The stability of this complex in the presence of high concentrations of serum and its high transduction efficiency in mice suggests that it is a promising candidate vehicle for in vivo gene delivery.

Published
2002

34. Poster Abstracts

Author

Littger, Ralf, primary, Alke, Alexandra, additional, Tewes, Bernhard, additional, Gropp, Felix, additional, Asai, T., additional, Watanabe, K., additional, Kuromi, K., additional, Kurohane, K., additional, Ogino, K., additional, Taki, T., additional, Tsukada, H., additional, Nakayama, J., additional, Oku, N., additional, Babai, I., additional, Matyas, G., additional, Baranji, L., additional, Milosevits, J., additional, Alving, C. R., additional, Bendas, G., additional, Rothe, U., additional, Scherphof, G. L., additional, Kamps, J. A. A. M., additional, Kessner, S., additional, Carafa, M., additional, Di Stefano, A., additional, Sozio, P., additional, Cacciatore, I., additional, Mosciatti, B., additional, Santucci, E., additional, Choice, E., additional, Harvie, P., additional, Galbraith, T., additional, Zunder, E., additional, Dutzar, B., additional, Anklesaria, P., additional, Paul, R., additional, Cocquyt, J., additional, De Cuyper, M., additional, Van der Meeren, P., additional, Cruz, M. E. M., additional, Gaspar, M. M., additional, Silva, M. T., additional, Dathe, M., additional, Nikolenko, H., additional, Wessolowski, A., additional, Schmieder, P., additional, Beyermann, M., additional, Bienert, M., additional, Santos, N. Dos, additional, Cox, K. A., additional, Allen, C., additional, Gallagher, R. C., additional, Ickenstein, L., additional, Mayer, L. D., additional, Bally, M. B., additional, Fischer, S., additional, Margalit, R., additional, Freisleben, H.-J., additional, Garidel, P., additional, Chen, H. C., additional, Moore, D., additional, Mendelsohn, R., additional, Keller, M., additional, Hildebrand, A., additional, Blume, A., additional, Girão da Cruz, M. T., additional, Simões, S., additional, Pedroso de Lima, M. C., additional, Graser, A., additional, Nahde, T., additional, Fahr, A., additional, Müller, R., additional, Müller-Brüsselbach, S., additional, Cudmore, S., additional, O'Mahony, D., additional, Hoving, S., additional, van Tiel, S. T., additional, Seynhaeve, A. L. B., additional, Ambagtsheer, G., additional, Eggermont, A. M. M., additional, ten Hagen, T. L. M., additional, Høyrup, P., additional, Jensen, S. S., additional, Jørgensen, K., additional, Iden, D., additional, Kuang, H., additional, Mullen, P., additional, Jacobs, C., additional, Roben, P., additional, Stevens, T., additional, Lollo, C., additional, Ishida, T., additional, Maeda, R., additional, Masuda, K., additional, Ichihara, M., additional, Kiwada, H., additional, Jung, K., additional, Reszka, R., additional, Kaiser, N., additional, Ohloff, I., additional, Linser-Haar, S., additional, Massing, U., additional, Schubert, R., additional, Kan, P., additional, Tsao, C. W., additional, Chen, W. K., additional, Wang, A. J., additional, Kimpfler, A., additional, Gerber, C., additional, Wieschollek, A., additional, Bruchelt, G., additional, Kobayashi, T., additional, Okada, Y., additional, Sone, S., additional, Harashima, H., additional, Maruyama, K., additional, Kondo, Masayo, additional, Lee, Chun Man, additional, Tanaka, Toshiyuki, additional, Su, Wei, additional, Kitagawa, Toru, additional, Ito, Toshinori, additional, Matsuda, Hikaru, additional, Murai, Toshiyuki, additional, Miyasaka, Masayuki, additional, Junji, Kimura, additional, Kondo, Masami, additional, Asai, Tomohiro, additional, Ogino, Koichi, additional, Taki, Takao, additional, Tsukada, Hideo, additional, Baba, Kazuhiko, additional, Oku, Naoto, additional, Koning, G. A., additional, Wauben, M. H. M., additional, Vestweber, D., additional, Everts, M., additional, Kok, R. J., additional, Schraa, A. J., additional, Molema, G., additional, Schiffelers, R. M., additional, Storm, G., additional, Kristl, J., additional, Šentjurc, M., additional, Abramović, Z., additional, Landry, S., additional, Perron, S., additional, Bestman-Smith, J., additional, Désormeaux, A., additional, Tremblay, M. J., additional, Bergeron, M. G., additional, Madeira, C., additional, Loura, L. M. S., additional, Fedorov, A., additional, Prieto, M., additional, Aires-Barros, M. R., additional, Marques, C. M., additional, Simões, S. I., additional, Cruz, M. E., additional, Cevc, G., additional, Martins, M. B., additional, Moreira, J. N., additional, Gaspar, R., additional, Allen, T. M., additional, Esposito, C., additional, Ortaggi, G., additional, Bianco, A., additional, Bonadies, F., additional, Malizia, D., additional, Napolitano, R., additional, Cametti, C., additional, Mossa, G., additional, Endert, Gerold, additional, Essler, Frank, additional, Lutz, Silke, additional, Panzner, Steffen, additional, Pastorino, F., additional, Brignole, C., additional, Pagnan, G., additional, Moase, E. H., additional, Ponzoni, M., additional, Pavelic, Z., additional, Škalko-Basnet, N., additional, Jalšenjak, I., additional, Penacho, N., additional, Pisano, C., additional, Bucci, F., additional, Serafini, S., additional, Martinelli, R., additional, Cupelli, A., additional, Marconi, A., additional, Ferrara, F. F., additional, Santaniello, M., additional, Critelli, L., additional, Tinti, O., additional, Luisi, P., additional, Carminati, P., additional, Galletti, B., additional, Sauer, I., additional, Schleef, M., additional, Voß, C., additional, Schmidt, T., additional, Flaschel, E., additional, König, S., additional, Wenger, T., additional, Dumond, J., additional, Bogetto, N., additional, Reboud-Ravaux, M., additional, Schramm, H. J., additional, Schramm, W., additional, Sheynis, T., additional, Rozner, S., additional, Kolusheva, S., additional, Satchell, D., additional, Jelnik, R., additional, Shigeta, Y., additional, Imanaka, H., additional, Ando, H., additional, Makino, T., additional, Baba, N., additional, Shimizu, K., additional, Takada, M., additional, Baba, K., additional, Namba, Y., additional, Simberg, Dmitri, additional, Danino, Dganit, additional, Talmon, Yeshayahu, additional, Minsky, Abraham, additional, Ferrari, Marilyn E., additional, Wheeler, Carl J., additional, Barenholz, Yechezkel, additional, Takada, Miki, additional, Shimizu, Kosuke, additional, Kuromi, Koici, additional, Takeuchi, Y., additional, North, J. R., additional, Nango, M., additional, Tewes, B., additional, Köchling, T., additional, Deissler, M., additional, Kühl, C., additional, Marx, U., additional, Strote, G., additional, Gropp, F., additional, Qualls, Marquita M., additional, Kim, Jong-Mok, additional, Thompson, David H., additional, Zhang, Zhi-Yi, additional, Shum, Pochi, additional, Collier, Joel H., additional, Hu, Bi-Huang, additional, Ruberti, Jeffrey W., additional, Messersmith, Phillip B., additional, Tsuruda, T., additional, Nakade, A., additional, Sadzuka, Y., additional, Hirota, S., additional, Sonobe, T., additional, Vorauer-Uhl, K., additional, Wagner, A., additional, Katinger, H., additional, Weeke-Klimp, A. H., additional, Bartsch, M., additional, Meijer, D. K. F., additional, Zeisig, R., additional, Walther, W., additional, Reß, A., additional, Fichtner, I., additional, Zschörnig, O., additional, Schiller, J., additional, Süß, M., additional, Bergmeier, C., additional, Arnold, K., additional, Nchinda, Godwin, additional, Überla, Klaus, additional, and Zschörnig, Olaf, additional

Published
2003
Full Text
View/download PDF

42. Poster Abstracts

Author

Littger, Ralf, Alke, Alexandra, Tewes, Bernhard, Gropp, Felix, Asai, T., Watanabe, K., Kuromi, K., Kurohane, K., Ogino, K., Taki, T., Tsukada, H., Nakayama, J., Oku, N., Babai, I., Matyas, G., Baranji, L., Milosevits, J., Alving, C. R., Bendas, G., Rothe, U., Scherphof, G. L., Kamps, J. A. A. M., Kessner, S., Rothe, U., Bendas, G., Carafa, M., Di Stefano, A., Sozio, P., Cacciatore, I., Mosciatti, B., Santucci, E., Choice, E., Harvie, P., Galbraith, T., Zunder, E., Dutzar, B., Anklesaria, P., Paul, R., Cocquyt, J., De Cuyper, M., Van der Meeren, P., Cruz, M. E. M., Gaspar, M. M., Silva, M. T., Dathe, M., Nikolenko, H., Wessolowski, A., Schmieder, P., Beyermann, M., Bienert, M., Santos, N. Dos, Cox, K. A., Allen, C., Gallagher, R. C., Ickenstein, L., Mayer, L. D., Bally, M. B., Fischer, S., Margalit, R., Freisleben, H.-J., Garidel, P., Chen, H. C., Moore, D., Mendelsohn, R., Garidel, P., Keller, M., Hildebrand, A., Blume, A., Girão da Cruz, M. T., Simões, S., Pedroso de Lima, M. C., Graser, A., Nahde, T., Fahr, A., Müller, R., Müller-Brüsselbach, S., Harvie, P., Dutzar, B., Choice, E., Cudmore, S., O'Mahony, D., Anklesaria, P., Paul, R., Hoving, S., van Tiel, S. T., Seynhaeve, A. L. B., Ambagtsheer, G., Eggermont, A. M. M., ten Hagen, T. L. M., Høyrup, P., Jensen, S. S., Jørgensen, K., Iden, D., Kuang, H., Mullen, P., Jacobs, C., Roben, P., Stevens, T., Lollo, C., Ishida, T., Maeda, R., Masuda, K., Ichihara, M., Kiwada, H., Jung, K., Reszka, R., Kaiser, N., Ohloff, I., Linser-Haar, S., Massing, U., Schubert, R., Kan, P., Tsao, C. W., Chen, W. K., Wang, A. J., Kimpfler, A., Gerber, C., Wieschollek, A., Bruchelt, G., Schubert, R., Kobayashi, T., Okada, Y., Ishida, T., Sone, S., Harashima, H., Maruyama, K., Kiwada, H., Kondo, Masayo, Lee, Chun Man, Tanaka, Toshiyuki, Su, Wei, Kitagawa, Toru, Ito, Toshinori, Matsuda, Hikaru, Murai, Toshiyuki, Miyasaka, Masayuki, Junji, Kimura, Kondo, Masami, Asai, Tomohiro, Ogino, Koichi, Taki, Takao, Tsukada, Hideo, Baba, Kazuhiko, Oku, Naoto, Koning, G. A., Wauben, M. H. M., ten Hagen, T. L. M., Vestweber, D., Everts, M., Kok, R. J., Schraa, A. J., Molema, G., Schiffelers, R. M., Storm, G., Kristl, J., Šentjurc, M., Abramovi, Z., Landry, S., Perron, S., Bestman-Smith, J., Désormeaux, A., Tremblay, M. J., Bergeron, M. G., Madeira, C., Loura, L. M. S., Fedorov, A., Prieto, M., Aires-Barros, M. R., Marques, C. M., Simões, S. I., Cruz, M. E., Cevc, G., Martins, M. B., Moreira, J. N., Gaspar, R., Allen, T. M., Esposito, C., Ortaggi, G., Bianco, A., Bonadies, F., Malizia, D., Napolitano, R., Cametti, C., Mossa, G., Endert, Gerold, Essler, Frank, Lutz, Silke, Panzner, Steffen, Pastorino, F., Brignole, C., Pagnan, G., Moase, E. H., Allen, T. M., Ponzoni, M., Pavelic, Z., Škalko-Basnet, N., Jalšenjak, I., Penacho, N., Simões, S., Pedroso de Lima, M. C., Pisano, C., Bucci, F., Serafini, S., Martinelli, R., Cupelli, A., Marconi, A., Ferrara, F. F., Santaniello, M., Critelli, L., Tinti, O., Luisi, P., Carminati, P., Santaniello, M., Bucci, F., Tinti, O., Pisano, C., Critelli, L., Galletti, B., Luisi, P., Carminati, P., Sauer, I., Nikolenko, H., Dathe, M., Schleef, M., Voß, C., Schmidt, T., Flaschel, E., König, S., Wenger, T., Dumond, J., Bogetto, N., Reboud-Ravaux, M., Schramm, H. J., Schramm, W., Sheynis, T., Rozner, S., Kolusheva, S., Satchell, D., Jelnik, R., Shigeta, Y., Imanaka, H., Ando, H., Makino, T., Kurohane, K., Oku, N., Baba, N., Shimizu, K., Asai, T., Takada, M., Baba, K., Namba, Y., Oku, N., Simberg, Dmitri, Danino, Dganit, Talmon, Yeshayahu, Minsky, Abraham, Ferrari, Marilyn E., Wheeler, Carl J., Barenholz, Yechezkel, Takada, Miki, Shimizu, Kosuke, Kuromi, Koici, Asai, Tomohiro, Baba, Kazuhiko, Oku, Naoto, Takeuchi, Y., Kurohane, K., North, J. R., Namba, Y., Nango, M., Oku, N., Tewes, B., Köchling, T., Deissler, M., Kühl, C., Marx, U., Strote, G., Gropp, F., Qualls, Marquita M., Kim, Jong-Mok, Thompson, David H., Zhang, Zhi-Yi, Shum, Pochi, Collier, Joel H., Hu, Bi-Huang, Ruberti, Jeffrey W., Messersmith, Phillip B., Thompson, David H., Tsuruda, T., Nakade, A., Sadzuka, Y., Hirota, S., Sonobe, T., Vorauer-Uhl, K., Wagner, A., Katinger, H., Wagner, A., Vorauer-Uhl, K., Katinger, H., Weeke-Klimp, A. H., Bartsch, M., Meijer, D. K. F., Scherphof, G. L., Kamps, J. A. A. M., Zeisig, R., Walther, W., Reß, A., Fichtner, I., Zschörnig, O., Schiller, J., Süß, M., Bergmeier, C., Arnold, K., Nchinda, Godwin, Überla, Klaus, and Zschörnig, Olaf

Abstract

DOCSPER—A Synthetic Lipid Fit for In Vivo ApplicationDOCSPER [1,3-Dioleoyloxy-2-(N5-carbamoyl-spermine)-propane] is a cationic amphiphile consisting of a hydrophobic 1,3 dioleylglycerol moiety and threefold positively charged spermine head group (1). We optimised the 5-step-synthesis of the lipospermine and after up-scaling we have obtained sufficient amounts to initiate preclinical investigations. DOCSPER was tested for its ability to transfect eukaryotic cells in vitro. It has proven to possess high transfection efficiency in comparison to commercially available liposomal transfection agents. Furthermore, DOCSPER was extensively tested in several in vivo studies (23). These studies revealed a high transfection efficiency, whereas very low toxicity levels were detected. Thus, the results clearly indicate that the cationic lipid DOCSPER is a reliable, low-risk system for broad applications in gene therapy.Groth D. et al. Int J Pharm 1998; 162:143–157.Nikol S. et al. Int J Angiol 2000; 9:87–95.Armeanu S. et al. Mol Ther 2000; 1(4):366–375.

Published
1982
Full Text
View/download PDF

44. Annexin A2/P11 interaction

Author

Schulz, Daniela M., Stefan Kalkhof, Schmidt, Andreas, Ihling, Christian, Stingl, Christoph, Mechtler, Karl, Zschörnig, Olaf, and Sinz, Andrea

45. Cholesterol is the major component of native lipoproteins activating the p38 mitogen-activated protein kinases

Author

Dobreva, Iveta, Zschörnig, Olaf, Waeber, Gérard, James, Richard W., Widmann, Christian, Dobreva, Iveta, Zschörnig, Olaf, Waeber, Gérard, James, Richard W., and Widmann, Christian

Abstract

Elevated low-density lipoprotein (LDL) levels induce activation of the p38 mitogen-activated protein kinase (MAPK), a stress-activated protein kinase potentially participating in the development of atherosclerosis. The nature of the lipoprotein components inducing p38 MAPK activation has remained unclear however. We show here that both LDLs and high-density lipoproteins (HDLs) have the ability to stimulate the p38 MAPKs with potencies that correlate with their cholesterol content. Cholesterol solubilized in methyl-β-cyclodextrin was sufficient to activate the p38 MAPK pathway. Liposomes made of phosphatidylcholine (PC) or sphingomyelin, the two main phospholipids found in lipoproteins, were unable to stimulate the p38 MAPKs. In contrast, PC liposomes loaded with cholesterol potently activated this pathway. Reducing the cholesterol content of LDL particles lowered their ability to activate the p38 MAPKs. Cell lines representative of the three main cell types found in blood vessels (endothelial cells, smooth muscle cells and fibroblasts) all activated their p38 MAPK pathway in response to LDLs or cholesterol-loaded PC liposomes. These results indicate that elevated cholesterol content in lipoproteins, as seen in hypercholesterolemia, favors the activation of the stress-activated p38 MAPK pathway in cells from the vessel wall, an event that might contribute to the development of atherosclerosis

48. Membrane binding and structure of de novo designed alpha-helical cationic coiled-coil-forming peptides.

Author

Vagt T, Zschörnig O, Huster D, and Koksch B

Subjects
Amino Acid Sequence, Biophysics methods, Carbon Isotopes chemistry, Chemistry, Physical methods, Circular Dichroism, Leucine chemistry, Lipids chemistry, Magnetic Resonance Spectroscopy, Membranes, Artificial, Molecular Conformation, Molecular Sequence Data, Phospholipids chemistry, Protein Folding, Protein Structure, Secondary, Peptides chemistry
Abstract

We introduce a de novo designed peptide model system that enables the systematic study of 1) the role of a membrane environment in coiled-coil peptide folding, 2) the impact of different domains of an alpha-helical coiled-coil heptad repeat on the interaction with membranes, and 3) the dynamics of coiled-coil peptide-membrane interactions depending on environmental conditions. Starting from an ideal alpha-helical coiled-coil peptide sequence, several positively charged analogues were designed that exhibit a high propensity toward negatively charged lipid membranes. Furthermore, these peptides differ in their ability to form a stable alpha-helical coiled-coil structure. The influence of a membrane environment on peptide folding is studied. All positively charged peptides show strong interactions with negatively charged membranes. This interaction induces an alpha-helical structure of the former random-coil peptides, as revealed by circular dichroism measurements. Furthermore, vesicle aggregation is induced by a coiled-coil interaction of vesicle-bound peptides. Dynamic light scattering experiments show that the strength of vesicle aggregation increases with the peptide's intrinsic ability to form a stable alpha-helical coiled coil. Thus, the peptide variant equipped with the strongest inter- and intra-helical coiled-coil interactions shows the strongest effect on vesicle aggregation. The secondary structure of this peptide in the membrane-bound state was studied as well as its effect on the phospholipids. Peptide conformation within the peptide-lipid aggregates was analyzed by (13)C cross-polarization magic-angle spinning NMR experiments. A uniformly (13)C- and (15)N-labeled Leu residue was introduced at position 12 of the peptide chain. The (13)C chemical shift and torsion angle measurements support the finding of an alpha-helical structure of the peptide in its membrane-bound state. Neither membrane leakage nor fusion was observed upon peptide binding, which is unusual for amphiphatic peptide structures. Our results lay the foundation for a systematic study of the influence of the alpha-helical coiled-coil folding motif in membrane-active events on a molecular level.

Published
2006
Full Text
View/download PDF

49. Structural determinants for plant annexin-membrane interactions.

Author

Dabitz N, Hu NJ, Yusof AM, Tranter N, Winter A, Daley M, Zschörnig O, Brisson A, and Hofmann A

Subjects
Animals, Annexins genetics, Annexins metabolism, Annexins ultrastructure, Capsicum chemistry, Gossypium chemistry, Membrane Lipids chemistry, Phospholipids chemistry, Phospholipids metabolism, Plant Proteins genetics, Plant Proteins metabolism, Plant Proteins ultrastructure, Protein Binding, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Annexins chemistry, Membrane Lipids metabolism, Plant Proteins chemistry
Abstract

The interactions of two plant annexins, annexin 24(Ca32) from Capsicum annuum and annexin Gh1 from Gossypium hirsutum, with phospholipid membranes have been characterized using liposome-based assays and adsorption to monolayers. These two plant annexins show a preference for phosphatidylserine-containing membranes and display a membrane binding behavior with a half-maximum calcium concentration in the sub-millimolar range. Surprisingly, the two plant annexins also display calcium-independent membrane binding at levels of 10-20% at neutral pH. This binding is regulated by three conserved surface-exposed residues on the convex side of the proteins that play a pivotal role in membrane binding. Due to quantitative differences in the membrane binding behavior of N-terminally His-tagged and wild-type annexin 24(Ca32), we conclude that the N-terminal domain of plant annexins plays an important role, reminiscent of the findings in their mammalian counterparts. Experiments elucidating plant annexin-mediated membrane aggregation and fusion, as well as the effect of these proteins on membrane surface hydrophobicity, agree with findings from the membrane binding experiments. Results from electron microscopy reveal elongated rodlike assemblies of plant annexins in the membrane-bound state. It is possible that these structures consist of protein molecules directly interacting with the membrane surface and molecules that are membrane-associated but not in direct contact with the phospholipids. The rodlike structures would also agree with the complex data from intrinsic protein fluorescence. The tubular lipid extensions suggest a role in the membrane cytoskeleton scaffolding or exocytotic processes. Overall, this study demonstrates the importance of subtle changes in an otherwise conserved annexin fold where these two plant annexins possess distinct modalities compared to mammalian and other nonplant annexins.

Published
2005
Full Text
View/download PDF

Catalog

Books, media, physical & digital resources
See catalog results