Home LiteratureArticle Details
PMID: 8779593 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Production of a polyhydroxyalkanoate biopolymer in insect cells with a modified eucaryotic fatty acid synthase.

Applied and environmental microbiology ·Vol. 62 ·No. 7 ·1996-07-00 ·Pages 2540-6

Williams MD, Rahn JA, Sherman DH

Abstract

A novel pathway for the synthesis of poly-3-hydroxybutyrate has been engineered by simultaneous delivery of two genes into insect cells (Spodoptera frugiperda) by use of individual baculovirus vectors. This system includes expression of a dehydrase-domain mutant rat fatty acid synthase cDNA and the phbC gene encoding polyhydroxyalkanoate synthase from Alcaligenes eutrophus. The dehydrase-deficient fatty acid synthase provides de novo synthesis of R-(-)-3-hydroxybutyryl-coenzyme A as a premature termination product rather than palmityl-coenzyme A, the normal product of wild-type rat fatty acid synthase. High levels of this mutant multifunctional protein provide a suitable precursor pool of R-(-)-3-hydroxybutyryl-coenzyme A for conversion to poly-3-hydroxybutyrate in insect cells coexpressing the phbC gene product. This strategy for redesigning a poly-3-hydroxybutyrate biosynthetic pathway suggests a new method for generating structurally diverse polyhydroxyalkanoates by metabolic engineering.

MeSH Terms
Animals Baculoviridae/genetics Biopolymers/biosynthesis,chemistry Cell Line Cytoplasmic Granules/metabolism DNA, Complementary/genetics Fatty Acid Synthases/genetics,metabolism Gene Expression Genetic Engineering Rats Spodoptera
Chemicals
Biopolymers DNA, Complementary Fatty Acid Synthases
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Williams M D
Department of Microbiology and Biological Process Technology Institute, St. Paul, Minnesota 55108-6106, USA.
Rahn J A
Sherman D H
References (36)
36 references, click to expand
  1. Presence of two polypeptide chains comprising fatty acid synthetase.
    Proc Natl Acad Sci U S A. 1975 May;72(5):1940-4 PMID: 1098047
  2. Cleavage of structural proteins during the assembly of the head of bacteriophage T4.
    Nature. 1970 Aug 15;227(5259):680-5 PMID: 5432063
  3. Construction, expression, and characterization of a mutated animal fatty acid synthase deficient in the dehydrase function.
    J Biol Chem. 1993 Oct 25;268(30):22508-13 PMID: 8226759
  4. Overexpression and purification of the soluble polyhydroxyalkanoate synthase from Alcaligenes eutrophus: evidence for a required posttranslational modification for catalytic activity.
    Biochemistry. 1994 Aug 9;33(31):9311-20 PMID: 8049232
  5. Isolation of a functional transferase component from the rat fatty acid synthase by limited trypsinization of the subunit monomer. Formation of a stable functional complex between transferase and acyl carrier protein domains.
    J Biol Chem. 1991 Oct 15;266(29):19180-5 PMID: 1918034
  6. Pseudomonas putida KT2442 cultivated on glucose accumulates poly(3-hydroxyalkanoates) consisting of saturated and unsaturated monomers.
    Appl Environ Microbiol. 1992 Feb;58(2):536-44 PMID: 1610179
  7. Bacterial polyesters containing branched poly(beta-hydroxyalkanoate) units.
    Int J Biol Macromol. 1990 Apr;12(2):92-101 PMID: 2078536
  8. STRUCTURE OF POLY-BETA-HYDROXYBUTYRIC ACID GRANULES.
    J Gen Microbiol. 1964 Mar;34:441-6 PMID: 14135548
  9. Isolation and mapping of the beta-hydroxyacyl dehydratase activity of chicken liver fatty acid synthase.
    J Biol Chem. 1988 Nov 5;263(31):16225-9 PMID: 3182791
  10. Physiology and molecular genetics of poly(beta-hydroxy-alkanoic acid) synthesis in Alcaligenes eutrophus.
    Mol Microbiol. 1991 Mar;5(3):535-42 PMID: 2046547
  11. Poly-beta-hydroxybutyrate (PHB) biosynthesis in Alcaligenes eutrophus H16. Identification and characterization of the PHB polymerase gene (phbC).
    J Biol Chem. 1989 Sep 15;264(26):15298-303 PMID: 2670936
  12. Poly-beta-hydroxybutyrate biosynthesis in Alcaligenes eutrophus H16. Characterization of the genes encoding beta-ketothiolase and acetoacetyl-CoA reductase.
    J Biol Chem. 1989 Sep 15;264(26):15293-7 PMID: 2670935
  13. Polyhydroxybutyrate, a biodegradable thermoplastic, produced in transgenic plants.
    Science. 1992 Apr 24;256(5056):520-3 PMID: 17787950
  14. Plastics from bacteria and for bacteria: poly(beta-hydroxyalkanoates) as natural, biocompatible, and biodegradable polyesters.
    Adv Biochem Eng Biotechnol. 1990;41:77-93 PMID: 2126418
  15. Occurrence, metabolism, metabolic role, and industrial uses of bacterial polyhydroxyalkanoates.
    Microbiol Rev. 1990 Dec;54(4):450-72 PMID: 2087222
  16. Targeting of the polyhydroxybutyrate biosynthetic pathway to the plastids of Arabidopsis thaliana results in high levels of polymer accumulation.
    Proc Natl Acad Sci U S A. 1994 Dec 20;91(26):12760-4 PMID: 7809117
  17. Expression and analysis of a bacterial poly(hydroxyalkanoate) synthase in insect cells using a baculovirus system.
    Protein Expr Purif. 1996 Mar;7(2):203-11 PMID: 8812862
  18. Polymeric Beta-Hydroxyalkanoates from Environmental Samples and Bacillus megaterium.
    Appl Environ Microbiol. 1983 Jan;45(1):71-8 PMID: 16346184
  19. Molecular genetics of polyketides and its comparison to fatty acid biosynthesis.
    Annu Rev Genet. 1990;24:37-66 PMID: 2088174
  20. Biosynthesis and characterization of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) in Alcaligenes eutrophus.
    Int J Biol Macromol. 1990 Apr;12(2):106-11 PMID: 2078526
  21. Production of polyhydroxyalkanoates, a family of biodegradable plastics and elastomers, in bacteria and plants.
    Biotechnology (N Y). 1995 Feb;13(2):142-50 PMID: 9634754
  22. Molecular cloning and sequencing of cDNAs encoding the entire rat fatty acid synthase.
    Proc Natl Acad Sci U S A. 1989 May;86(9):3114-8 PMID: 2717611
  23. Production of unsaturated polyesters by Pseudomonas oleovorans.
    Int J Biol Macromol. 1990 Apr;12(2):85-91 PMID: 2078535
  24. Specific release of the thioesterase component of the fatty acid synthetase multienzyme complex by limited trypsinization.
    Proc Natl Acad Sci U S A. 1976 Apr;73(4):1184-8 PMID: 1063400
  25. Modular structure of genes encoding multifunctional peptide synthetases required for non-ribosomal peptide synthesis.
    FEMS Microbiol Lett. 1995 Jan 1;125(1):3-14 PMID: 7867917
  26. The role and regulation of energy reserve polymers in micro-organisms.
    Adv Microb Physiol. 1973;10:135-266 PMID: 4594739
  27. Branched-chain fatty acids produced by mutants of Streptomyces fradiae, putative precursors of the lactone ring of tylosin.
    Antimicrob Agents Chemother. 1990 Aug;34(8):1535-41 PMID: 2221862
  28. Repositioning of a domain in a modular polyketide synthase to promote specific chain cleavage.
    Science. 1995 Jun 9;268(5216):1487-9 PMID: 7770773
  29. Assembly of five bluetongue virus proteins expressed by recombinant baculoviruses: inclusion of the largest protein VP1 in the core and virus-like proteins.
    Virology. 1991 Feb;180(2):798-802 PMID: 1846500
  30. The animal fatty acid synthase: one gene, one polypeptide, seven enzymes.
    FASEB J. 1994 Dec;8(15):1248-59 PMID: 8001737
  31. Recovery of Poly-beta-Hydroxybutyrate from Estuarine Microflora.
    Appl Environ Microbiol. 1978 Feb;35(2):251-7 PMID: 16345267
  32. Multifunctional proteins.
    Annu Rev Biochem. 1976;45:143-66 PMID: 786148
  33. Studies on the mechanism of fatty acid synthesis. X. Reduced triphosphopyridine nucleotide-acetoacetyl coenzyme A reductase.
    J Biol Chem. 1962 Mar;237:687-93 PMID: 14004466
  34. Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure.
    Proc Natl Acad Sci U S A. 1987 Nov;84(21):7413-7 PMID: 2823261
  35. Malonyl and palmityl transferase-less mutants of the yeast fatty-acid-synthetase complex.
    Eur J Biochem. 1975 Aug 15;56(2):359-67 PMID: 1100391
  36. Construction of a cDNA encoding the multifunctional animal fatty acid synthase and expression in Spodoptera frugiperda cells using baculoviral vectors.
    Biochem J. 1993 Nov 15;296 ( Pt 1):143-9 PMID: 8250834
Article Info
Journal
Applied and environmental microbiology
Abbr.
Appl Environ Microbiol
ISSN
0099-2240
DOI
10.1128/aem.62.7.2540-2546.1996
Published
1996-07-00
Pages
2540-6
Language
English
Region
United States
NLM ID
7605801
PMCID
PMC168036
Subset
IM
Grants
NIDDK NIH HHS · DK16073 · United States
Corrections
ErratumIn
-
External Links
PubMed source DOI: 10.1128/aem.62.7.2540-2546.1996 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2