doi:10.1016/j.biotechadv.2006.11.007
Copyright © 2006 Elsevier Inc. All rights reserved.
Biotechnological approaches for the production of polyhydroxyalkanoates in microorganisms and plants — A review
References and further reading may be available for this article. To view references and further reading you must
purchase this article.
Pornpa Suriyamongkola, b, Randall Weselakeb, Suresh Narineb, Maurice Moloneyc and Saleh Shaha, 
, 
aPlant Biotechnology Unit, Alberta Research Council, Vegreville, Alberta, Canada T9C 1T4
bDepartment of Agricultural, Food and Nutritional Sciences, University of Alberta, Edmonton, Alberta, Canada T6G 2P5
cDepartment of Biological Sciences, University of Calgary, Calgary, Canada T2N 1N4
Received 14 August 2006;
revised 23 November 2006;
accepted 23 November 2006.
Available online 30 November 2006.
Abstract
The increasing effect of non-degradable plastic wastes is a growing concern. Polyhydroxyalkanoates (PHAs), macromolecule-polyesters naturally produced by many species of microorganisms, are being considered as a replacement for conventional plastics. Unlike petroleum-derived plastics that take several decades to degrade, PHAs can be completely bio-degraded within a year by a variety of microorganisms. This biodegradation results in carbon dioxide and water, which return to the environment. Attempts based on various methods have been undertaken for mass production of PHAs. Promising strategies involve genetic engineering of microorganisms and plants to introduce production pathways. This challenge requires the expression of several genes along with optimization of PHA synthesis in the host. Although excellent progress has been made in recombinant hosts, the barriers to obtaining high quantities of PHA at low cost still remain to be solved. The commercially viable production of PHA in crops, however, appears to be a realistic goal for the future.
Keywords: Polyhydroxyalkanoates; PHA; Polyhydroxybutyrate; PHB; Bioplastics; Microorganisms; E. coli; Yeast; Transgenic plants
Abbreviations: ACP, acyl carrier protein; CaMV35S, cauliflower mosaic virus 35S; CoA, coenzyme A; DGAT, diacylglycerol acyltransferase; dwt, dry weight; ER, endoplasmic reticulum; fad, fatty acid desaturase; HA, 3-hydroxyalkanoate; HB, 3-hydroxybutyrate; 4HB, 4-hydroxybutyrate; HD, 3-hydroxydecanoate; HH, 3-hydroxyhexanoate; HO, 3-hydroxyoctanoate; HV, 3-hydroxyvalerate; lcl-, long-chain-length; mcl-, medium-chain-length; PCR, polymerase chain reaction; PHA, polyhydroxyalkanoate; PhaA, β-ketothiolase; PhaB, acetoacetyl-CoA reductase; PhaC, PHA synthase; PHB, poly(3-hydroxybutyrate); P(HB-HH), poly(3-hydroxybutyrate-co-3-hydroxyhexanoate); P(HB-HV), poly(3-hydroxybutyrate-co-3-hydroxyvalerate); PDC, pyruvate dehydrogenase complex; scl-, short-chain-length.
Fig. 1. Chemical structure of PHAs. The pendant R groups (shaded boxes) vary in chain length from one carbon (C1) to over 14 carbons (C14). Structures shown here are PHB (R = methyl), PHV (R = ethyl), and PHH (R = propyl).
Fig. 2. PHB and P(HB–HV) biosynthetic pathways in R. eutropha. phaA and bktB encode β-ketothiolase and 3-ketothiolase, enzymes involved in formation of acetoacetyl-CoA and 3-ketovaleryl-CoA, respectively. phaB encodes acetoacetyl-CoA reductase, which reduces both acetoacetyl-CoA and 3-ketovaleryl-CoA to form (R)-3-hydroxybutyryl-CoA and (R)-3-hydroxyvaleryl-CoA, respectively. phaC encodes PHA synthase, which is the last enzyme responsible for polymerization of the monomers (adapted from Poirier, 2002).
Fig. 3. pha gene operons in different microorganisms. (a) A complete phaCAB operon of R. eutropha; (b) interrupted locus of Z. ramigera; (c) locus with two polymerase subunits, phaC and phaE, of C. vinosum; (d) phaCJ operon of A. caviae for copolymer P(HB–HH) formation; (e) phaC1ZC2D operon for mcl-PHA formation in P. oleovorans with two phaC genes (C1 and C2); (f) pha locus with depolymerase (phaZ) between two polymerase subunits in P. aureofaciens and phaF gene situated downstream of phaC1ZC2D operon.
Fig. 4. Fatty acid β-oxidation pathway of E. coli. Recombinant E. coli with defective fadA and fadB uses intermediates of β-oxidation, enoyl-CoA, 3-ketoacyl-CoA and (S)-3-hydroxyacyl-CoA, as major substrates for mcl-PHA synthesis. Co-expression of mcl-PHA synthase with enoyl-CoA hydratase (encoded by paaF, paaG, ydbU, maoC, yfcX from E. coli; phaJ from P. aeruginosa, P. putida and A. caviae) or 3-ketoacyl-CoA/ACP reductase (encoded by fabG from P. aeruginosa and E. coli; rhlG from P. aeruginosa) can enhance PHA production by increasing β-oxidation intermediate pools (adapted from Park and Lee, 2003).
Fig. 5. Fatty acid de novo biosynthesis (Fab pathway). P. aeruginosa and P. putida use (R)-3-hydroxyacyl-CoA monomers from Fab pathway to produce mcl-PHAs through expression of PhaG and PhaC when grown on carbon sources which is metabolized to acetyl-CoA, like carbohydrate. PhaG acts as a link between Fab pathway and mcl-PHA synthesis by catalyzing (R)-3-hydroxyacyl-ACP, intermediate of Fab pathway, to (R)-3-hydroxyacyl-CoA, substrate for mcl-PHA.
Fig. 6. Genetically engineered metabolic pathways for PHB synthesis in plant cytoplasm. Dashed arrows indicate expression of transgenes, while plant native pathways are shown by solid arrows. Enhancement of acetyl-CoA pool by enzyme inhibitors is also shown. Crosses indicate targeted sites of enzyme inhibitors. Quizalofop inhibits acetyl-CoA carboxylase (ACCase) of the flavonoid synthetic pathway and mevastatin inhibits 3-hydroxy-3-methylglutaryl-CoA (HMGR) of the mevalonic synthetic pathway.
Fig. 7. Various plant transformation gene constructs. (a) Constructs of phaB and phaC from R. eutropha under constitutive CaMV35S (35S) promoters with Hygromycin (HPT) and Kanamycin (NPTII) resistance; (b) three individual plastid target constructs with each gene attached to the chloroplast transit peptide (CTP) and under CaMV35S promoter; (c) triple plastid target construct with CaMV35S promoter and chloroplast transit peptide; (d) polycistronic mRNA pha operon from R. eutropha for chloroplast genomic transformation under prrn promoter; (e) peroxisomal target construct of phaC1 from P. aeruginosa for mcl-PHA formation with the attachment of DNA segment encoding the last 34 amino acids of B. napus isocitrate lyase (ICL). Tnos, terminator sequence from nopaline synthase gene; TpsbA, terminator sequence from plastid psbA gene; RB, right border of the T-DNA; LB, left border of the T-DNA.
Fig. 8. Genetically engineered PHB and copolymer P(HB–HV) synthetic pathways in plant plastids. ilvA from E. coli encodes threonine deaminase. phaA, bktB, phaB, and phaC are from R. eutropha. phaA and bktB encode thiolases with different substrate specificities; phaB and phaC encode acetoacetyl-CoA reductase and PHA synthase, respectively. PDC refers to pyruvate dehydrogenase complex in plant plastids. The cross indicates the targeted site of enzyme inhibitor, Quizalofop, which enhances the acetyl-CoA pool in the engineered pathway. Dashed arrows indicate expression of transgenes, while plant native pathways are shown in solid arrows.
Fig. 9. Genetically engineered metabolic pathways of scl- and mcl-PHA formation in plant peroxisomes. Expressions of transgenes are indicated by dashed arrows. phaC from A. caviae and P. aeruginosa encode PHA synthases that use (R)-3-hydroxyacyl-CoA from fatty acid degradation as substrate for scl- and mcl-PHA polymerization, respectively.
Fig. 10. Strategies of synthesizing PHA in peroxisomes with special monomers, using lipid biosynthesis mutant (e.g. DGAT mutant); co-expressing fatty acid biosynthetic genes (e.g. acyl-ACP hydrolase); exogenous feeding of special fatty acids, etc. (adapted from Poirier, 2002).
Corresponding author. Fax: +1 780 632 8612.
Abstract
Purchase PDF (938 K)
E-mail Article
Add to my Quick Links
Cited By in Scopus (8)
Purchase PDF (168 K)
