PMID:15757895
| Citation |
Lüer, C, Schauer, S, Möbius, K, Schulze, J, Schubert, WD, Heinz, DW, Jahn, D and Moser, J (2005) Complex formation between glutamyl-tRNA reductase and glutamate-1-semialdehyde 2,1-aminomutase in Escherichia coli during the initial reactions of porphyrin biosynthesis. J. Biol. Chem. 280:18568-72 |
|---|---|
| Abstract |
In Escherichia coli the first common precursor of all tetrapyrroles, 5-aminolevulinic acid, is synthesized from glutamyl-tRNA (Glu-tRNA(Glu)) in a two-step reaction catalyzed by glutamyl-tRNA reductase (GluTR) and glutamate-1-semialdehyde 2,1-aminomutase (GSA-AM). To protect the highly reactive reaction intermediate glutamate-1-semialdehyde (GSA), a tight complex between these two enzymes was proposed based on their solved crystal structures. The existence of this hypothetical complex was verified by two independent biochemical techniques. Co-immunoprecipitation experiments using antibodies directed against E. coli GluTR and GSA-AM demonstrated the physical interaction of both enzymes in E. coli cell-free extracts and between the recombinant purified enzymes. Additionally, the formation of a GluTR.GSA-AM complex was identified by gel permeation chromatography. Complex formation was found independent of Glu-tRNA(Glu) and cofactors. The analysis of a GluTR mutant truncated in the 80-amino acid C-terminal dimerization domain (GluTR-A338Stop) revealed the importance of GluTR dimerization for complex formation. The in silico model of the E. coli GluTR.GSA-AM complex suggested direct metabolic channeling between both enzymes to protect the reactive aldehyde species GSA. In accordance with this proposal, side product formation catalyzed by GluTR was observed via high performance liquid chromatography analysis in the absence of the GluTR.GSA-AM complex. |
| Links |
PubMed Online version:10.1074/jbc.M500440200 |
| Keywords |
Aldehyde Oxidoreductases/chemistry; Aldehydes/chemistry; Blotting, Western; Catalysis; Cell-Free System; Chromatography, Gel; Chromatography, High Pressure Liquid; Crystallography, X-Ray; Dimerization; Escherichia coli/chemistry; Escherichia coli/enzymology; Escherichia coli/metabolism; Gene Deletion; Glutamates/chemistry; Immunoblotting; Immunoprecipitation; Intramolecular Transferases/chemistry; Models, Molecular; Mutagenesis, Site-Directed; Mutation; Porphyrins/chemistry; Protein Structure, Tertiary; Recombinant Proteins/chemistry; Time Factors |
| edit table |
Significance
You can help EcoliWiki by summarizing why this paper is useful
Useful Materials and Methods
You can help Ecoliwiki by describing the useful materials (strains, plasmids, antibodies, etc) described in this paper.
Annotations
<annotationlinks/>
EcoliWiki Links
Add links to pages that link here (e.g. gene, product, method pages)
References
See Help:References for how to manage references in EcoliWiki.