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Cloning and sequence analysis of the poly(3-hydroxyalkanoic acid)-synthesis genes ofPseudomonas acidophila

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Abstract

Pseudomonas acidophila can grow with CO2 as a sole carbon source by the possession of a recombinant plasmid that clones genes that confer chemolithoautotrophic growth ability derived from the H2-oxidizing bacteriumAlcaligenes hydrogenophilus. H2-oxidizing bacteria produce poly(3-hydroxybutyric acid) (PHB) from CO2, but recombinant P.acidophila can produce the more useful biopolymer poly(3-hydroxyalkanoic acid) (PHA). In this study, thepha genes ofP. acidophila were cloned and a sequence analysis was carried out. A gene library was constructed using the cosmid vector pVK102. A recombinant cosmid carrying thepha genes was selected by the complementation of a PHB-negative mutant ofAlcaligenes eutrophus H16. The resulting recombinant cosmid pIK7 contained a 14.8-kb DNA insert. Subcloning was done, and the recombinant plasmid pEH74 was selected by hybridization with theA. eutrophus H16pha genes.Escherichia coli possessing pEH74 produced PHB, indicating that pEH74 contained thepha genes ofP. acidophila. The nucleotide sequences of the PHA-synthesis genesphaA (3-ketothiolase),phaB (acetoacetyl-CoA reductase), andphaC (PHA synthase) in pEH74 were determined. The homologies ofphaA, phaB, andphaC betweenP. acidophila andA. eutrophus H16 were 64.7, 76.1, and 56.6%, respectively.

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Authors and Affiliations

  1. Faculty of Pharmaceutical Sciences, Osaka University, 1-6 Yamada-oka, Suita, 565, Osaka, Japan

    Fusako Umeda, Yoshiharu Kitano, Yuki Murakami, Kiyohito Yagi, Yoshiharu Miura & Tadashi Mizoguchi

Authors
  1. Fusako Umeda
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  2. Yoshiharu Kitano
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  3. Yuki Murakami
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  4. Kiyohito Yagi
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  5. Yoshiharu Miura
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  6. Tadashi Mizoguchi
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Umeda, F., Kitano, Y., Murakami, Y. et al. Cloning and sequence analysis of the poly(3-hydroxyalkanoic acid)-synthesis genes ofPseudomonas acidophila . Appl Biochem Biotechnol 70, 341–352 (1998). https://doi.org/10.1007/BF02920150

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