Biosynthesis of Polylactic Acid and Its Copolymers Using Evolved Propionate CoA Transferase and PHA Synthase

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For the synthesis of polylactic acid (PLA) and its copolymers by one-step fermentation process, heterologous pathways involving Clostridium propionicum propionate CoA transferase (Pct(Cp)) and Pseudomonas sp. MBEL 6-19 polyhydroxyalkanoate (PHA) synthase I (PhaC1(Ps6-19)) were introduced into Escherichia coli for the generation of lactyl-CoA endogenously and incorporation of lactyl-CoA into the polymer, respectively. Since the wild-type PhaC1(Ps6-19) did not efficiently accept lactyl-CoA as a substrate, site directed mutagenesis as well as saturation mutagenesis were performed to improve the enzyme. The wild-type Pct(CP) was not able to efficiently convert lactate to lactyl-CoA and was found to exert inhibitory effect on cell growth, random mutagenesis by error-prone PCR was carried out. By employing engineered PhaC1(Ps6-19) and Pct(Cp), poly(3-hydroxybutyrate-co-lactate), P(3HB-co-LA), containing 20-49mol% lactate could be produced up to 62wt% from glucose and 3HB. By controlling the 3HB concentration in the medium, PLA homopolymer and P(3HB-co-LA) containing lactate as a major monomer unit could be synthesized. Also, P(3HB-co-LA) copolymers containing various lactate fractions could be produced from glucose alone by introducing the Cupriavidus necator beta-ketothiolase and acetoacetyl-CoA reductase genes. Fed-batch cultures were performed to produce P(3HB-co-LA) copolymers having 9-64 mol% of lactate, and their molecular weights, thermal properties, and melt flow properties were determined. Biotechnol. Bioeng. 2010;105: 150-160. (C) 2009 Wiley Periodicals, Inc.
Publisher
JOHN WILEY SONS INC
Issue Date
2010-01
Language
English
Article Type
Article
Keywords

RECOMBINANT ESCHERICHIA-COLI; POLYHYDROXYALKANOATE SYNTHASES; SUBSTRATE-SPECIFICITY; POLY(LACTIC ACID); MOLECULAR-WEIGHT; IN-VIVO; CLONING; POLY(3-HYDROXYBUTYRATE); POLYESTERS; ENZYME

Citation

BIOTECHNOLOGY AND BIOENGINEERING, v.105, no.1, pp.150 - 160

ISSN
0006-3592
DOI
10.1002/bit.22547
URI
http://hdl.handle.net/10203/20255
Appears in Collection
CBE-Journal Papers(저널논문)
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