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Cloning and sequencing of the hemA gene of Rhodobacter capsulatus and isolation of a δ-aminolevulinic acid-dependent mutant strain

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Summary

The Rhodobacter capsulatus hemA gene, coding for the enzyme δ-aminolevulinic acid synthase (ALAS), was isolated from a genome bank by hybridization with a hemT probe from Rhodobacter sphaeroides. Subcloning of the initial 3.9 kb HindIII fragment allowed the isolation of a 2.5 kb HindIII-BglII fragment which was able to complement the δ-aminolevulinic acid-requiring (ALA-requiring) Escherichia coli mutant SHSP19. DNA sequencing revealed an open reading frame coding for a protein with 401 amino acids which displayed similarity to the amino acid sequences of other known ALASs. However, no resemblance was seen to the HemA protein of E. coli K12. Based on the sequence data, an ALA-requiring mutant strain of R. capsulatus was constructed by site-directed insertion mutagenesis. Introduction of a plasmid, containing the hemA gene of R. capsulatus on the 3.9 kb HindIII fragment, restored ALA-independent growth of the mutant indicating that there is only one gene for ALA biosynthesis in R. capsulatus. Transfer of the R′ factor pRPS404 and hybridization analysis revealed that the ALAS gene is not located within the major photosynthetic gene cluster.

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Authors and Affiliations

  1. Institut für Biologie II, Mikrobiologie, Universität Freiburg, Schänzlestrasse 1, D-7800, Freiburg, Germany

    Ulrike Hornberger, Rainer Liebetanz, Hans-Volker Tichy & Gerhart Drews

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  1. Ulrike Hornberger
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  2. Rainer Liebetanz
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  3. Hans-Volker Tichy
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  4. Gerhart Drews
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Communicated by H. Hennecke

Part of this research was presented at the Symposium on Molecular Biology of Membrane-Bound Complexes in Phototrophic Bacteria, Freiburg, FRG, 2–5 August 1989

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Hornberger, U., Liebetanz, R., Tichy, HV. et al. Cloning and sequencing of the hemA gene of Rhodobacter capsulatus and isolation of a δ-aminolevulinic acid-dependent mutant strain. Mol Gen Genet 221, 371–378 (1990). https://doi.org/10.1007/BF00259402

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  • DOI: https://doi.org/10.1007/BF00259402

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