Current Microbiology

, Volume 50, Issue 6, pp 329–333 | Cite as

Genetic Characterization of the Poly(hydroxyalkanoate) Synthases of Various Pseudomonas oleovorans Strains

  • Daniel K.Y. SolaimanEmail author
  • Richard D. Ashby


We identified the poly(hydroxyalkanoate) synthase (PHAS) genes of three strains of Pseudomonas oleovorans by using polymerase chain reaction (PCR)-based detection methods. P. oleovorans NRRL B-14682 contains Class I PHA synthase gene (phaC), NRRL B-14683 harbors Class II phaC1 and phaC2 genes, and NRRL B-778 contain both the Class I and II PHA synthase genes. Inverse-PCR and chromosomal walking techniques were employed to obtain the complete sequences of the Class I phaCs of NRRL B-778 (phbC778; 1698 bps) and B-14682 (phbC14682; 1899 bps). BLAST search indicated that these genes are new and had not been previously cloned. The gene product of phbC778 (i.e., PhbC778; 566 amino acid residues) is homologous to the Class I PHA synthases of Pseudomonas sp. HJ-2 and Pseudomonas sp. strain 61-3, and that of phbC14682 (PhbC14682; 632 amino acids) is homologous to PHAS of Delftia acidovorans. The PhbC14682 contains an extra sequence of 33 amino acids in its conserved α/β-hydrolase domain, making it only the second Class I PHA synthase found to contain this cellular proteolytic sequence. Consistent with their Pseudomonas origin, the codon-usage profiles of PhbC778 and PhbC14682 are similar to those of Pseudomonas Class II PHASs. These new Pseudomonas Class I phbC genes provide valuable addition to the gene pool for the construction of novel PHASs through gene shuffling.


Polymerase Chain Reaction Method Extra Sequence Polymerase Chain Reaction Procedure Pseudomonas Oleovorans PHAS Gene 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



The technical assistance of Nicole Cross and Marshall Reed is acknowledged.

Literature Cited

  1. 1.
    Aldor, IS, Keasling, JD 2003Process design for microbial plastic factories: metabolic engineering of polyhydroxyalkanoatesCurr Opin Biotechnol14475483PubMedGoogle Scholar
  2. 2.
    Altschul, SF, Madden, TL, Schäffer, AA, Zhang, J, Zhang, Z, Miller, W, Lipman, DJ 1997Gapped BLAST and PSI-BLAST: a new generation of protein database search programsNucleic Acids Res2533893402CrossRefPubMedGoogle Scholar
  3. 3.
    Ashby, RD, Solaiman, DKY, Foglia, TA 2002aThe synthesis of short- and medium-chain-length poly(hydroxyalkanoate) mixtures from glucose- or alkanoic acid-grown Pseudomonas oleovoransJ Ind Microbiol Biotechnol28147153Google Scholar
  4. 4.
    Ashby, RD, Solaiman, DKY, Foglia, TA 2002bPoly(ethylene glycol)-mediated molar mass control of short-chain- and medium-chain-length poly(hydroxyalkanoates) from Pseudomonas oleovoransAppl Microbiol Biotechnol60154159Google Scholar
  5. 5.
    Besse, P, Veschambre, H 1994Chemical and biological synthesis of chiral epoxidesTetrahedron5088858927Google Scholar
  6. 6.
    Chenna, R, Sugawara, H, Koike, T, Lopez, R, Gibson, TJ, Higgins, DG, Thompson, JD 2003Multiple sequence alignment with the Clustal series of programsNucleic Acids Res3134973500PubMedGoogle Scholar
  7. 7.
    Hazenberg, W, Witholt, B 1997Efficient production of medium-chain-length poly(3-hydroxyalkanoates) from octane by Pseudomonas oleovorans: economic considerationsAppl Microbiol Biotechnol48588596Google Scholar
  8. 8.
    Huisman, GW, Wonink, E, Meima, R, Kazemier, B, Terpstra, P, Witholt, B 1991Metabolism of poly(3-hydroxyalkanoates) (PHAs) by Pseudomonas oleovorans. Identification and sequences of genes and function of the encoded proteins in the synthesis and degradation of PHAJ Biol Chem26621912198PubMedGoogle Scholar
  9. 9.
    Rehm, BHA 2003Polyester synthases: natural catalysts for plasticsBiochem J3761533PubMedGoogle Scholar
  10. 10.
    Sambrook, J, Russell, DW 2001Molecular cloning: a laboratory manual, 3rd ed, vol. 2Cold Spring Harbor Laboratory PressCold Spring Harbor, NY881885Google Scholar
  11. 11.
    Sheu, D-S, Wang, Y-T, Lee, C-Y 2000Rapid detection of polyhydroxyalkanoate-accumulating bacteria isolated from the environment by colony PCRMicrobiology14620192025PubMedGoogle Scholar
  12. 12.
    Solaiman, DKY 2002Polymerase-chain-reaction-based detection of individual polyhydroxyalkanoate synthase phaC1 and phaC2 genesBiotechnol Lett24245250Google Scholar
  13. 13.
    Solaiman, DKY 2003Biosynthesis of medium-chain-length poly(hydroxyalkanoates) with altered composition by mutant hybrid PHA synthasesJ Ind Microbiol Biotechnol30322326PubMedGoogle Scholar
  14. 14.
    Solaiman, DKY, Ashby, RD, Foglia, TA 2000Rapid and specific identification of medium-chain-length polyhydroxyalkanoate synthase gene by polymerase chain reactionAppl Microbiol Biotechnol53690694PubMedGoogle Scholar
  15. 15.
    Steinbüchel, A, Lütke-Eversloh, T 2003Metabolic engineering and pathway construction for biotechnological production of relevant polyhydroxyalkanoates in microorganismsBiochem Eng J168196Google Scholar
  16. 16.
    Sudesh, K, Fukui, T, Doi, Y 1998Genetic analysis of Comamonas acidovorans polyhydroxyalkanoate synthase and factors affecting the incorporation of 4-hydroxybutyrate monomerAppl Environ Microbiol6434373443PubMedGoogle Scholar
  17. 17.
    Tatusova, TA, Madden, TL 1999Blast 2 sequences - a new tool for comparing protein and nucleotide sequencesFEMS Microbiol Lett174247250PubMedGoogle Scholar
  18. 18.
    Tsuge, T, Imazua, S-i, Takaseb, K, Taguchib, S, Doi, Y 2004An extra large insertion in the polyhydroxyalkanoate synthase from Delftia acidovorans DS-17: its deletion effects and relation to cellular proteolysisFEMS Microbiol Lett2317783PubMedGoogle Scholar
  19. 19.
    Beilen, JB, Eggink, G, Enequist, H, Bos, R, Witholt, B 1992DNA sequence determination and functional characterization of the OCT-plasmid-encoded alkJKL genes of Pseudomonas oleovoransMol Microbiol631213136PubMedGoogle Scholar
  20. 20.
    Beilen, JB, Panke, S, Lucchini, S, Franchini, AG, Röthlisberger, M, Witholt, B 2001Analysis of Pseudomonas putida alkane-degradation gene clusters and flanking insertion sequences: evolution and regulation of the alk genesMicrobiology14716211630PubMedGoogle Scholar
  21. 21.
    Witholt, B, Kessler, B 1999Perspectives of medium chain length poly(hydroxyalkanoates), a versatile set of bacterial bioplasticsCurr Opin Biotechnol10279285PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, Inc. 2005

Authors and Affiliations

  1. 1.Fats, Oils and Animal Coproducts Research Unit, Eastern Regional Research Center, Agricultural Research ServiceUnited States Department of AgricultureWyndmoorUSA

Personalised recommendations