Abstract
Four (R)-specific enoyl CoA hydratases (PhaJ) interconnect the β-oxidation pathway with PHA biosynthesis in Pseudomonas aeruginosa. The use of antisense technique and over-expression to delineate the role of two of these enzymes, PhaJ1 and PhaJ4 forms the basis of this study. It has been observed that P. aeruginosa recombinant with phaJ1 antisense construct, fed with different fatty acids, produces PHA with less hydroxy octanoate (7–11% reduction) and a proportionate increase in other monomer fractions, compared to that of the control. Recombinants bearing phaJ4 antisense construct are found to contain less hydroxy decanoate (10–11% reduction) and more or less equal amount of hydroxy octanoate, compared to that of the control. P. aeruginosa has produced PHA with more hydroxy octanoate and decanoate (6–17% increase), respectively, when PhaJ1 and PhaJ4 have been over-expressed individually or along with PhaC1. PhaJ1 and PhaJ4 are found to be involved mainly in the production of hydroxy octanoyl CoA and hydroxy decanoyl CoA, respectively, in P. aeruginosa. The strongest accumulation of hydroxy octanoate and hydroxy decanoate has been observed along with hydroxy butyrate, in PHA, produced by E. coli, when PhaC1 has been co-expressed with PhaJ1 and PhaJ4, respectively. We have demonstrated, for the first time, the polymerization of hydroxy butyryl CoA monomers in recombinant E. coli by PhaC1 of P. aeruginosa.
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Acknowledgements
The authors wish to thank Department of Biotechnology, India, for funding the project. Reeta Davis gratefully acknowledges Council of Scientific and Industrial Research, India, for research fellowship. The LS1298Kan::fadB1 was kindly provided by Eliza McKinney of CLF Medical Technology Acceleration Program, Inc. New York, USA. We are grateful for Herbert P. Schweizer of Colorado State University for having provided the plasmid pBSP II KS (−) and the strain PA01.We thank Mr. P.S. Kulashekhar, C.F.T.R.I. for editing the manuscript.
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Davis, R., Chandrashekar, A. & Shamala, T.R. Role of (R)-specific enoyl coenzyme A hydratases of Pseudomonas sp in the production of polyhydroxyalkanoates. Antonie van Leeuwenhoek 93, 285–296 (2008). https://doi.org/10.1007/s10482-007-9203-1
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DOI: https://doi.org/10.1007/s10482-007-9203-1