Genetic "Carbohydrate-Engineering" of S-layer glycoproteins of Gram-positive eubacteria (P14209-MOB)
Abstract
The goal of the proposed research project is the production for the first time, of a bacterial S-layer glycoprotein which comprises an artificial glycan chain altered by molecular genetic methods. In addition, we propose to lay the groundwork for the subsequent design of genetically engineered bacterial glycoproteins comprising the glycan chains of capsular polysaccharides or mammalian proteoglycans. These are important target structures for use as innovative vaccines and biocompatible matrices. This work is scheduled for a duration of 3 years. The Gram-positive, thermophilic eubacteria Aneurinibacillus thermoaerophilus L420-91T and GS4-97, as well as Bacillus stearothermophilus NRS 2004/3a have been chosen as model organisms. The aim of the proposal is the specific modification of the S-layer glycoprotein glycan chain from A. thermoaerophilus by "carbohydrate engineering" to create an artificial S-layer neoglycoprotein. Such experiments using prokaryotic glycoproteins have not been pursued so far. As an important prerequisite, cloning and sequencing of the genes involved in the biosynthesis of GDP-D-rhamnose should be accomplished within the first year. In year two, the genes encoding the synthesis of the amino sugar should be cloned and sequenced. In year three, the glycan structures of the S-layer glycoproteins of A. thermoaerophilus will be modified by insertional mutagenesis. For specific applications of S-layer glycoproteins, the formation of uniform surface structures is of great importance. Therefore, the enzymes catalyzing the polymerization of the glycan repeating units will also be investigated in year three. The proposed investigations are expected to provide important foundations for the production of specifically designed S-layer neoglycoproteins useful in biomedical and nanotechnological applications.
keywords Carbohydrate-Engineering S-layers bacterial glycoprotein glycosylation enzymes glycosylation genes
Publikationen
Synthesis of GDP-D-rhamnose from GDP-D-mannose in Aneurinibacillus thermoaerophilus L420-91.
Autoren: Kneidinger, B., Graninger, M., Messner, P. Jahr: 2000
Conference & Workshop proceedings, paper, abstract
A pyrophosphate bridge links the pyruvate-containing secondary cell wall polymer of Paenibacillus alvei CCM 2051 to muramic acid.
Autoren: Schäffer, C; Müller, N; Mandal, PK; Christian, R; Zayni, S; Messner, P Jahr: 2000
Journal articles
Chromosome walking by cloning of distinct PCR fragments.
Autoren: Kneidinger, B; Graninger, M; Messner, P Jahr: 2001
Journal articles
Identification of two GDP-6-deoxy-D-lyxo-4-hexulose reductases synthesizing GDP-D-rhamnose in Aneurinibacillus thermoaerophilus L420-91T.
Autoren: Kneidinger, B; Graninger, M; Adam, G; Puchberger, M; Kosma, P; Zayni, S; Messner, P Jahr: 2001
Journal articles
The crystal structure of dTDP-D-Glucose 4,6-dehydratase (RmlB) from Salmonella enterica serovar Typhimurium, the second enzyme in the dTDP-l-rhamnose pathway.
Autoren: Allard, ST; Giraud, MF; Whitfield, C; Graninger, M; Messner, P; Naismith, JH Jahr: 2001
Journal articles
Genetisches "Carbohydrate-Engineering" von S-Schichtglykoproteinen von Gram-positiven Eubakterien.
Autoren: Messner, P. Jahr: 2001
Forschungsbericht (extern. Auftraggeber)
Cloning and functional characterization of the Pseudomonas aeruginosa rhlC gene that encodes rhamnosyltransferase 2, an enzyme responsible for di-rhamnolipid biosynthesis.
Autoren: Rahim, R; Ochsner, UA; Olvera, C; Graninger, M; Messner, P; Lam, JS; Soberón-Chávez, G Jahr: 2001
Journal articles
Biosynthesis of nucleotide-activated D-glycero-D-manno-heptose.
Autoren: Kneidinger, B; Graninger, M; Puchberger, M; Kosma, P; Messner, P Jahr: 2001
Journal articles
Molecular characterization of the glycosylation of S-layer glycoproteins from Gram-positive bacteria.
Autoren: Kneidinger, B., Schäffer, C., Graninger, M., Pföstl, A., Novotny, R., Schröfelbauer, B., Sleytr, U.B., Messner, P. Jahr: 2001
Conference & Workshop proceedings, paper, abstract
Biosynthesis of nucleotide-activated D-glycero-D-manno-heptose: a phosphatase as missing link.
Autoren: Kneidinger, B., Graninger, M., Messner, P. Jahr: 2001
Conference & Workshop proceedings, paper, abstract
Glycobiology of surface layer proteins.
Autoren: Schäffer, C; Messner, P Jahr: 2001
Journal articles
Molecular characterization of S-layer glycoproteins from Gram-positive bacteria.
Autoren: Kneidinger, B., Graninger, M., Novotny, R., Pföstl, A., Puchberger, M., Zayni, S., Scheberl, A., Kosma, P., Schäffer, C., Sleytr, U.B., Messner, P. Jahr: 2001
Conference & Workshop proceedings, paper, abstract
S-layer glycoproteins and secondary cell wall polymers: major glycosylated wall components of Gram-positive bacteria.
Autoren: Schäffer, C., Kählig, H., Christian, R., Müller, N., Scheberl, A., Zayni, S., Sleytr, U.B., Messner, P. Jahr: 2001
Conference & Workshop proceedings, paper, abstract
Mapping and investigation of the cardiolipins in Bacillus stearothermophilus NRS 2004/3a by positive and negative ion nano ESI-QTOF-MS and MS/MS.
Autoren: Beckedorf, A.I., Schäffer, C., Messner, P., Peter-Katalinic, J. Jahr: 2001
Conference & Workshop proceedings, paper, abstract
Mapping and investigation of the cardiolipins in Bacillus stearothermophilus NRS 2004/3a by positive and negative ion nano ESI-QTOF-MS and MS/MS.
Autoren: Beckedorf, A.I., Schäffer, C., Messner, P., Peter-Katalinic, J. Jahr: 2001
Conference & Workshop proceedings, paper, abstract
A model for the biosynthesis of the S-layer glycoprotein of Aneurinibacillus thermoaerophilus DSM 10155.
Autoren: Graninger, M., Kneidinger, B., Puchberger, M., Kosma, P., Schäffer, C., Messner, P. Jahr: 2001
Conference & Workshop proceedings, paper, abstract
Biosynthesis pathway of ADP-L-glycero-beta-D-manno-heptose in Escherichia coli.
Autoren: Kneidinger, B; Marolda, C; Graninger, M; Zamyatina, A; McArthur, F; Kosma, P; Valvano, MA; Messner, P Jahr: 2002
Journal articles
Toward a structural understanding of the dehydratase mechanism.
Autoren: Allard, ST; Beis, K; Giraud, MF; Hegeman, AD; Gross, JW; Wilmouth, RC; Whitfield, C; Graninger, M; Messner, P; Allen, AG; Maskell, DJ; Naismith, JH Jahr: 2002
Journal articles
The surface layer (S-layer) glycoprotein of Geobacillus stearothermophilus NRS 2004/3a. Analysis of its glycosylation.
Autoren: Schäffer, C; Wugeditsch, T; Kählig, H; Scheberl, A; Zayni, S; Messner, P; Jahr: 2002
Journal articles
S-layer glycoproteins and S-layer neoglycoproteins.
Autoren: Novotny, R., Pföstl, A., Messner, P., Sleytr, U.B., Schäffer, C. Jahr: 2002
Conference & Workshop proceedings, paper, abstract
N-Acetyl-D-fucosamine biosynthesis in Aneurinibacillus thermoaerophilus L420-91.
Autoren: Pföstl, A., Hofinger, A., Novotny, R., Kosma, P., Messner, P. Jahr: 2002
Conference & Workshop proceedings, paper, abstract
Variation on a theme of SDR. dTDP-6-deoxy-L- lyxo-4-hexulose reductase (RmlD) shows a new Mg2+-dependent dimerization mode.
Autoren: Blankenfeldt, W; Kerr, ID; Giraud, MF; McMiken, HJ; Leonard, G; Whitfield, C; Messner, P; Graninger, M; Naismith, JH Jahr: 2002
Journal articles
Screening of bacterial lipid libraries by MALDI-TOF PSD and MALDI-QTOF CID.
Autoren: Beckedorf, A.I., Jovanovic, M., Fischer, W., Schäffer, C., Messner, P., Peter-Katalinic, J. Jahr: 2002
Conference & Workshop proceedings, paper, abstract
Novel pathways for biosynthesis of nucleotide-activated glycero-manno-heptose precursors of bacterial glycoproteins and cell surface polysaccharides.
Autoren: Valvano, MA; Messner, P; Kosma, P Jahr: 2002
Journal articles
Homologs of the Rml enzymes from Salmonella enterica are responsible for dTDP-beta-L-rhamnose biosynthesis in the gram-positive thermophile Aneurinibacillus thermoaerophilus DSM 10155.
Autoren: Graninger, M; Kneidinger, B; Bruno, K; Scheberl, A; Messner, P Jahr: 2002
Journal articles
Mapping and sequencing of cardiolipins from Geobacillus stearothermophilus NRS 2004/3a by positive and negative ion nanoESI-QTOF-MS and MS/MS.
Autoren: Beckedorf, AI; Schäffer, C; Messner, P; Peter-Katalinic, J Jahr: 2002
Journal articles
Functional expression of enterobacterial O-polysaccharide biosynthesis enzymes in Bacillus subtilis.
Autoren: Schäffer, C; Wugeditsch, T; Messner, P; Whitfield, C; Jahr: 2002
Journal articles
Glycosylation of S-layer proteins from Gram-positive bacteria
Autoren: Novotny, R., Pföstl, A., Schäffer, C., Zayni, S., Scheberl, A., Sleytr, U.B., Messner, P. Jahr: 2002
Conference & Workshop proceedings, paper, abstract
NMR-Strukturanalyse des sekundären Zellwandpolymers von Aneurinibacillus thermoaerophilus DSM 10155
Autoren: Steindl, C., Schäffer, C., Messner, P., Müller, N. Jahr: 2002
Conference & Workshop proceedings, paper, abstract
Chemical synthesis and biosynthetic pathways of nucleotide-activated heptoses
Autoren: Kneidinger, B., Zamyatina, A., Valvano, M., Messner, P., Kosma, P. Jahr: 2002
Conference & Workshop proceedings, paper, abstract
Isolation of glucocardiolipins from Geobacillus stearothermophilus NRS 2004/3a.
Autoren: Schäffer, C; Beckedorf, AI; Scheberl, A; Zayni, S; Peter-Katalinić, J; Messner, P; Jahr: 2002
Journal articles
Prokaryotic Glycoproteins
Autoren: Schäffer, C., Messner, P. Jahr: 2003
Chapter in collected volumes
Glycosylation of Bacterial Surface Layer (S-layer) Proteins
Autoren: Schäffer, C., Novotny, R., Pföstl, A., Sleytr, U.B., Messner, P. Jahr: 2003
Conference & Workshop proceedings, paper, abstract
The first biantennary bacterial secondary cell wall polymer and its influence on S-layer glycoprotein assembly.
Autoren: Steindl, C; Schäffer, C; Wugeditsch, T; Graninger, M; Matecko, I; Müller, N; Messner, P Jahr: 2002
Journal articles
Novel aspects of S-layer protein glycosilation
Autoren: Schäffer, C., Pföstl, A., Novotny, R., Giry-Laterriere, M., Steiner, K., Scheberl, A., Zayni, S., Sleytr, U.B., Messner, P. Jahr: 2003
Conference & Workshop proceedings, paper, abstract
The first biantennary bacterial secondary cell wall polymer and its influence on S-layer glycoprotein assembly
Autoren: Schäffer, C., Steindl, C., Pföstl, A., Müller, N., Sleytr, U.B., Messner, P. Jahr: 2003
Conference & Workshop proceedings, paper, abstract
Formation of nucleotide-activated sugar precursors required for the biosynthesis of S-layer glycoproteins
Autoren: Pföstl, A., Novotny, R., Zayni, S., Scheberl, A., Schäffer, C., Sleytr, U.B., Messner, P. Jahr: 2003
Conference & Workshop proceedings, paper, abstract
Biosynthesis of dTDP-3-acetamido-3,6-dideoxy-alpha-D-galactose in Aneurinibacillus thermoaerophilus L420-91T.
Autoren: Pfoestl, A; Hofinger, A; Kosma, P; Messner, P Jahr: 2003
Journal articles
Surface-layer glycoproteins: an example for the diversity of bacterial glycosylation with promising impacts on nanobiotechnology.
Autoren: Schäffer, C; Messner, P Jahr: 2004
Journal articles
S-layer glycan-specific loci on the chromosome of Geobacillus stearothermophilus NRS 2004/3a and dTDP-L-rhamnose biosynthesis potential of G. stearothermophilus strains.
Autoren: Novotny, R; Schäffer, C; Strauss, J; Messner, P Jahr: 2004
Journal articles
Genetic organization of chromosomal S-layer glycan biosynthesis loci of Bacillaceae.
Autoren: Novotny, R; Pfoestl, A; Messner, P; Schäffer, C Jahr: 2004
Journal articles
Gene cloning, functional expression and secretion of the S-layer protein SgsE from Geobacillus stearothermophilus NRS 2004/3a in Lactococcus lactis.
Autoren: Novotny, R; Scheberl, A; Giry-Laterriere, M; Messner, P; Schäffer, C Jahr: 2005
Journal articles
Project staff
Paul Messner
Ao.Univ.Prof.i.R. Dipl.-Ing. Dr.nat.techn. Paul Messner
paul.messner@boku.ac.at
Project Leader
01.05.2000 - 30.04.2003
Christina Schäffer
Univ.Prof. Dipl.-Ing. Dr.nat.techn. Christina Schäffer
christina.schaeffer@boku.ac.at
Tel: +43 1 47654-80203
Project Staff
01.05.2000 - 30.04.2003
BOKU partners
External partners
University of Guelph
Prof.Chris WHITFIELD, Prof.Joseph S. LAM
partner
University of Western Ontario
Prof. Dr. Miguel A. Valvano
partner
University of Copenhagen, Faculty of Health Sciences, School of Dentistry
Prof. Dr. Henrik Clausen
partner
The University St. Andrews, Centre for Biomolecular Sciences
Prof. Dr. James H. Naismith
partner