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dc.contributor.authorAntonio, RV
dc.contributor.authorSteinbuchel, A
dc.contributor.authorRehm, BHA
dc.date.accessioned2018-04-20T04:07:46Z
dc.date.available2018-04-20T04:07:46Z
dc.date.issued2000
dc.identifier.issn0378-1097
dc.identifier.doi10.1016/S0378-1097(99)00578-9
dc.identifier.urihttp://hdl.handle.net/10072/373510
dc.description.abstractIn order to investigate the in vivo substrate specificity of the type I polyhydroxyalkanoate (PHA) synthase from Ralstonia eutropha, we functionally expressed the PHA synthase gene in various Escherichia coli mutants affected in fatty acid β-oxidation and the wild-type. The PHA synthase gene was expressed either solely (pBHR70) or in addition to the R. eutropha genes encoding β-ketothiolase and acetoacetyl-coenzyme A (CoA) reductase comprising the entire PHB operon (pBHR68) as well as in combination with the phaC1 gene (pBHR77) from Pseudomonas aeruginosa encoding type II PHA synthase. The fatty acid β-oxidation route was employed to provide various 3-hydroxyacyl-CoA thioesters, depending on the carbon source, as in vivo substrate for the PHA synthase. In vivo PHA synthase activity was indicated by PHA accumulation and substrate specificity was revealed by analysis of the comonomer composition of the respective polyester. Only in recombinant E. coli fad mutants harboring plasmid pBHR68, the R. eutropha PHA synthase led to accumulation of poly(3-hydroxybutyrate-co-3-hydroxyoctanoate) (poly(3HB-co-3HO)) and poly(3HB-co-3HO-co-3-hydroxydodecanoate (3HDD)), when octanoate and decanoate or dodecanoate were provided as carbon source, respectively. Coexpression of phaC1 from P. aeruginosa indicated and confirmed the provision of PHA precursor via the β-oxidation pathway and led to the accumulation of a blend of two different PHAs in the respective E. coli strain. These data strongly suggested that R. eutropha PHA synthase accepts, besides the main substrate 3-hydroxybutyryl-CoA, also the CoA thioesters of 3HO and 3HDD.
dc.description.peerreviewedYes
dc.languageEnglish
dc.publisherElsevier Science
dc.relation.ispartofpagefrom111
dc.relation.ispartofpageto117
dc.relation.ispartofissue1
dc.relation.ispartofjournalFEMS Microbiology Letters
dc.relation.ispartofvolume182
dc.subject.fieldofresearchMicrobiology not elsewhere classified
dc.subject.fieldofresearchBiological Sciences
dc.subject.fieldofresearchAgricultural and Veterinary Sciences
dc.subject.fieldofresearchMedical and Health Sciences
dc.subject.fieldofresearchcode060599
dc.subject.fieldofresearchcode06
dc.subject.fieldofresearchcode07
dc.subject.fieldofresearchcode11
dc.titleAnalysis of in vivo substrate specificity of the PHA synthase from Ralstonia eutropha: Formation of novel copolyesters in recombinant Escherichia coli
dc.typeJournal article
dc.type.descriptionC1 - Articles
dc.type.codeC - Journal Articles
gro.hasfulltextNo Full Text
gro.griffith.authorRehm, Bernd


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