We report the results of cloning genes for two key biosynthetic enzymes of different 5-aminolevulinic acid (ALA) biosynthetic routes from Streptomyces. The genes encode the glutamyl-tRNAGlu reductase (GluTR) of the C5 pathway and the ALA synthase (ALAS) of the Shemin pathway. While Streptomyces coelicolor A3(2) synthesizes ALA via the C5 route, both pathways are operational in Streptomyces nodosus subsp. asukaensis, a producer of asukamycin. In this strain, the C5 route produces ALA for tetrapyrrole biosynthesis; the ALA formed by the Shemin pathway serves as a precursor of the 2-amino-3-hydroxycyclopent-2-enone moiety (C5N unit), an antibiotic component. The growth of S. nodosus and S. coelicolor strains deficient in the GluTR genes (gtr) is strictly dependent on ALA or heme supplementation, whereas the defect in the ALAS-encoding gene (hemA-asuA) abolishes the asukamycin production in S. nodosus. The recombinant hemA-asuA gene was expressed in Escherichia coli and in Streptomyces, and the encoded enzyme activity was demonstrated both in vivo and in vitro. The hemA-asuA gene is situated within a putative cluster of asukamycin biosynthetic genes. This is the first report about the cloning of genes for two different ALA biosynthetic routes from a single bacterium.

• MeSH
• 5-Aminolevulinate Synthetase genetics   metabolism   MeSH
• RNA, Transfer, Amino Acyl metabolism   MeSH
• DNA Primers MeSH
• Kinetics MeSH
• Cloning, Molecular MeSH
• Aminolevulinic Acid metabolism   MeSH
• Plasmids MeSH
• Polyenes metabolism   MeSH
• Recombinant Proteins metabolism   MeSH
• Restriction Mapping MeSH
• Streptomyces genetics   metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 5-Aminolevulinate Synthetase MeSH
• RNA, Transfer, Amino Acyl MeSH
• asukamycin MeSH Browser
• DNA Primers MeSH
• glutamyl-tRNA(Gln) MeSH Browser
• Aminolevulinic Acid MeSH
• Polyenes MeSH
• Recombinant Proteins MeSH

Southern hybridization with probes designed for detection of WD-repeats coding sequences gave positive results in 21 streptomycete strains indicating that WD-repeats encoding genes are massively spread among streptomycetes. One of them, the wdlA gene of Streptomyces lincolnensis, codes for a 971 amino acid protein with seven WD-repeats in its C-terminus, two transmembrane domains and an ATP/GTP binding site upstream of the WD-repeat region.

• MeSH
• ATP-Binding Cassette Transporters genetics   MeSH
• Bacterial Proteins genetics   metabolism   MeSH
• DNA Probes MeSH
• Cloning, Molecular MeSH
• Membrane Proteins genetics   MeSH
• Molecular Sequence Data MeSH
• GTP-Binding Proteins genetics   MeSH
• Repetitive Sequences, Amino Acid genetics   physiology   MeSH
• Amino Acid Sequence MeSH
• Sequence Analysis, DNA MeSH
• Sequence Alignment MeSH
• Blotting, Southern MeSH
• Streptomyces genetics   growth & development   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• ATP-Binding Cassette Transporters MeSH
• Bacterial Proteins MeSH
• DNA Probes MeSH
• Membrane Proteins MeSH
• GTP-Binding Proteins MeSH
• wdlA protein, Streptomyces lincolnensis MeSH Browser

The gene pkwA coding for a typical WD-repeat protein was found in the chromosome of the bacterium Thermomonospora curvata CCM 3352. Until now WD-repeat proteins were through to be confined to eukaryotes.

• MeSH
• Actinomycetales enzymology   genetics   MeSH
• Genes, Bacterial MeSH
• Bacterial Proteins genetics   MeSH
• Molecular Sequence Data MeSH
• Protein Serine-Threonine Kinases genetics   MeSH
• Repetitive Sequences, Nucleic Acid * MeSH
• Amino Acid Sequence MeSH
• Base Sequence MeSH
• Sequence Homology, Amino Acid MeSH
• Publication type
• Journal Article MeSH
• Comparative Study MeSH
• Names of Substances
• Bacterial Proteins MeSH
• PkwA protein, Thermomonospora curvata MeSH Browser
• Protein Serine-Threonine Kinases MeSH

DNA-Dependent RNA polymerase (EC 2.7.7.6) was isolated from Thermomonospora curvata. The purification steps included precipitation with Polymin P, elution of the precipitate with 0.3 mol/L KCl, precipitation with ammonium sulfate, affinity chromatography on heparin-agarose and molecular filtration on Biogel A 1.5 m.

• MeSH
• Actinomycetales enzymology   MeSH
• Bacterial Proteins isolation & purification   MeSH
• Chemical Precipitation MeSH
• Chromatography, Affinity MeSH
• DNA-Directed RNA Polymerases isolation & purification   MeSH
• Chromatography, Gel MeSH
• Molecular Weight MeSH
• Spores, Bacterial MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Bacterial Proteins MeSH
• DNA-Directed RNA Polymerases MeSH