title: Chimeric enzyme composed of polyhydroxyalkanoate (PHA) synthases from Ralstonia eutropha and Aeromonas caviae enhances production of PHAs in recombinant Escherichia coli.
abstract: Chimeric enzymes composed of polyhydroxyalkanoate (PHA) synthases from Ralstonia eutropha (Cupriavidus necator) (PhaC(Re)) and Aeromonas caviae (PhaC(Ac)) were constructed. PhaC(Re) is known for its potent enzymatic activity among the characterized PHA synthases. PhaCAc has broad substrate specificity and synthesizes short-chain-length (SCL)/medium-chain-length (MCL) PHA. We attempted to create chimeric enzymes inheriting both of the advantageous properties. Among eight chimeras, AcRe12, with 26% of the N-terminal of PhaC(Ac) and 74% of the C-terminal of PhaC(Re), exhibited comparable P(3-hydroxybutyrate) accumulation as parental enzymes in Escherichia coli JM109. Thus, AcRe12 was applied to SCL/MCL PHA production using E. coli LS5218 as the host. AcRe12 accumulated higher amount of PHA (50 wt %) than the parental enzymes. Furthermore, the PHA consisted of 2 mol % 3-hydroxyhexanoate as well as 3-hydroxybutyrate. Therefore, the chimeric PHA synthase, AcRe12, inherited the character of both of the parental enzymes and thus exhibits improved enzymatic properties.
authors.affiliation: Division of Biotechnology and Macromolecular Chemistry, Graduate School of Engineering, Hokkaido University, N13-W8, Kita-ku, Sapporo 060-8628, Japan.
authors.forename: Ken'ichiro
authors.initials: K
authors.lastname: Matsumoto
authors.forename: Kazuma
authors.initials: K
authors.lastname: Takase
authors.forename: Yoko
authors.initials: Y
authors.lastname: Yamamoto
authors.forename: Yoshiharu
authors.initials: Y
authors.lastname: Doi
authors.forename: Seiichi
authors.initials: S
authors.lastname: Taguchi
mesh_terms: D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism*; D000333:Aeromonas / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D020118:Cupriavidus necator / Q000201:enzymology* / Q000235:genetics; D017931:DNA Primers; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005818:Genetic Engineering; D015151:Immunoblotting; D008969:Molecular Sequence Data; D054813:Polyhydroxyalkanoates / Q000378:metabolism*; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D017931:DNA Primers; D004274:DNA, Recombinant; D054813:Polyhydroxyalkanoates; D011993:Recombinant Fusion Proteins; D000217:Acyltransferases; C077225:poly(3-hydroxyalkanoic acid) synthase