Search

Your search keyword '"Christina Schäffer"' showing total 2 results
Start Over Author "Christina Schäffer" Journal biochemical journal
2 results on '"Christina Schäffer"'

1. Biosynthesis of dTDP-3-acetamido-3,6-dideoxy-α-<scp>D</scp>-glucose

Author

Paul Messner, Sonja Zayni, Christina Schäffer, Andreas Pföstl, Andreas Hofinger, and Paul Kosma

Subjects
DNA, Bacterial, Glycan, Isomerase, medicine.disease_cause, Polymerase Chain Reaction, Biochemistry, Catalysis, Article, Substrate Specificity, 03 medical and health sciences, chemistry.chemical_compound, Biosynthesis, Deoxy Sugars, Escherichia coli, medicine, Thymine Nucleotides, Cloning, Molecular, Nuclear Magnetic Resonance, Biomolecular, Molecular Biology, Chromatography, High Pressure Liquid, Thermoanaerobacterium thermosaccharolyticum, DNA Primers, 030304 developmental biology, Thermophilic organism, chemistry.chemical_classification, 0303 health sciences, Base Sequence, biology, 030306 microbiology, Acetylation, Cell Biology, biology.organism_classification, Enzymes, Enzyme, chemistry, Dehydratase, biology.protein, Thermoanaerobacterium
Abstract

Derivatives of 3-amino-3,6-dideoxyhexoses are widespread in Nature. They are part of the repeating units of lipopolysaccharide O-antigens, of the glycan moiety of S-layer (bacterial cell surface layer) glycoproteins and also of many antibiotics. In the present study, we focused on the elucidation of the biosynthesis pathway of dTDP-alpha-D-Quip3NAc (dTDP-3-acetamido-3,6-dideoxy-alpha-D-glucose) from the Gram-positive, anaerobic, thermophilic organism Thermoanaerobacterium thermosaccharolyticum E207-71, which carries Quip3NAc in its S-layer glycan. The biosynthesis of dTDP-alpha-D-Quip3NAc involves five enzymes, namely a transferase, a dehydratase, an isomerase, a transaminase and a transacetylase, and follows a pathway similar to that of dTDP-alpha-D-Fucp3NAc (dTDP-3-acetamido-3,6-dideoxy-alpha-D-galactose) biosynthesis in Aneurinibacillus thermoaerophilus L420-91(T). The ORFs (open reading frames) of interest were cloned, overexpressed in Escherichia coli and purified. To elucidate the enzymatic cascade, the different products were purified by HPLC and characterized by NMR spectroscopy. The initiating reactions catalysed by the glucose-1-phosphate thymidylyltransferase RmlA and the dTDP-D-glucose-4,6-dehydratase RmlB are well established. The subsequent isomerase was shown to be capable of forming a dTDP-3-oxo-6-deoxy-D-glucose intermediate from the RmlB product dTDP-4-oxo-6-deoxy-D-glucose, whereas the isomerase involved in the dTDP-alpha-D-Fucp3NAc pathway synthesizes dTDP-3-oxo-6-deoxy-D-galactose. The subsequent reaction steps of either pathway involve a transaminase and a transacetylase, leading to the specific production of nucleotide-activated 3-acetamido-3,6-dideoxy-alpha-D-glucose and 3-acetamido-3,6-dideoxy-alpha-D-galactose respectively. Sequence comparison of the ORFs responsible for the biosynthesis of dTDP-alpha-D-Quip3NAc revealed homologues in Gram-negative as well as in antibiotic-producing Gram-positive bacteria. There is strong evidence that the elucidated biosynthesis pathway may also be valid for LPS (lipopolysaccharide) O-antigen structures and antibiotic precursors.

Published
2008

2. Biosynthesis of dTDP-3-acetamido-3,6-dideoxy-α-D-glucose.

Author

Andreas Pföstl, Sonja Zayni, Andreas Hofinger, Paul Kosma, Christina Schäffer, and Paul Messner

Subjects
BIOSYNTHESIS, GRAM-positive bacteria, ANAEROBIC bacteria, THERMOPHILIC microorganisms, ENZYMES, GLUCOSE
Abstract

Derivatives of 3-amino-3,6-dideoxyhexoses are widespread in Nature. They are part of the repeating units of lipopolysaccharide O-antigens, of the glycan moiety of S-layer (bacterial cell surface layer) glycoproteins and also of many antibiotics. In the present study, we focused on the elucidation of the biosynthesis pathway of dTDP-α-D-Quip3NAc (dTDP-3-acetamido-3,6-dideoxy-α-D-glucose) from the Gram-positive, anaerobic, thermophilic organism Thermoanaerobacterium thermosaccharolyticum E207-71, which carries Quip3NAc in its S-layer glycan. The biosynthesis of dTDP-α-D-Quip3NAc involves five enzymes, namely a transferase, a dehydratase, an isomerase, a transaminase and a transacetylase, and follows a pathway similar to that of dTDP-α-D-Fucp3NAc (dTDP-3-acetamido-3,6-dideoxy-α-D-galactose) biosynthesis in Aneurinibacillus thermoaerophilus L420-91T. The ORFs (open reading frames) of interest were cloned, overexpressed in Escherichia coli and purified. To elucidate the enzymatic cascade, the different products were purified by HPLC and characterized by NMR spectroscopy. The initiating reactions catalysed by the glucose-1-phosphate thymidylyltransferase RmlA and the dTDP-D-glucose-4,6-dehydratase RmlB are well established. The subsequent isomerase was shown to be capable of forming a dTDP-3-oxo-6-deoxy-D-glucose intermediate from the RmlB product dTDP-4-oxo-6-deoxy-D-glucose, whereas the isomerase involved in the dTDP-α-D-Fucp3NAc pathway synthesizes dTDP-3-oxo-6-deoxy-D-galactose. The subsequent reaction steps of either pathway involve a transaminase and a transacetylase, leading to the specific production of nucleotide-activated 3-acetamido-3,6-dideoxy-α-D-glucose and 3-acetamido-3,6-dideoxy-α-D-galactose respectively. Sequence comparison of the ORFs responsible for the biosynthesis of dTDP-α-D-Quip3NAc revealed homologues in Gram-negative as well as in antibiotic-producing Gram-positive bacteria. There is strong evidence that the elucidated biosynthesis pathway may also be valid for LPS (lipopolysaccharide) O-antigen structures and antibiotic precursors. [ABSTRACT FROM AUTHOR]

Published
2008
Full Text
View/download PDF

Catalog

Books, media, physical & digital resources
See catalog results