Linked Life Data

11120644

Source:http://linkedlifedata.com/resource/pubmed/id/11120644

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
4
pubmed:dateCreated
2001-2-13
pubmed:abstractText
Multicopy plasmids are often chosen for the expression of recombinant genes in Escherichia coli. The high copy number is generally desired for maximum gene expression; however, the metabolic burden effects that usually result from multiple plasmid copies could prove to be detrimental for maximum productivity in certain metabolic engineering applications. In this study, low-copy mini-F plasmids were compared to high-copy pMB1-based plasmids for production of two metabolites in E. coli: polyphosphate (polyP) and lycopene derived from isopentenyl diphosphate (IPP). The stationary-phase accumulation of polyP on a per cell basis was enhanced approximately 80% when either high- or low-copy plasmids were used, from 120 micromol/g DCW without augmented polyP kinase (PPK) activity to approximately 220 micromol/g DCW. The cell density of the high-copy plasmid-containing culture at stationary phase was approximately 24% lower than the low-copy culture and 30% lower than the control culture. This difference in cell density is likely a metabolic burden effect and resulted in a lower overall product concentration for the high-copy culture (approximately 130 micromol/L culture) relative to the low-copy culture (approximately 160 micromol/L culture). When the gene for DXP (1-deoxy-D-xylulose 5-phosphate) synthase, the first enzyme in the IPP mevalonate-independent biosynthetic pathway, was expressed from the tac promoter on multicopy and low-copy plasmids, lycopene production was enhanced two- to threefold over that found in cells expressing the chromosomal copy only. Cell growth and lycopene production decreased substantially when isopropyl beta-D-thiogalactosidase (IPTG) was added to the high-copy plasmid-containing culture, suggesting that overexpression of DXP synthase was a significant metabolic burden. In the low-copy plasmid-containing culture, no differences in cell growth or lycopene production were observed with any IPTG concentrations. When dxs was placed under the control of the arabinose-inducible promoter (P(BAD)) on the low-copy plasmid, the amount of lycopene produced was proportional to the arabinose concentration and no significant changes in cell growth resulted. These results suggest that low-copy plasmids may be useful in metabolic engineering applications, particularly when one or more of the substrates used in the recombinant pathway are required for normal cellular metabolism.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
http://linkedlifedata.com/resource/pubmed/chemical/Acid Anhydride Hydrolases, http://linkedlifedata.com/resource/pubmed/chemical/Arabinose, http://linkedlifedata.com/resource/pubmed/chemical/Carotenoids, http://linkedlifedata.com/resource/pubmed/chemical/Hemiterpenes, http://linkedlifedata.com/resource/pubmed/chemical/Isopropyl Thiogalactoside, http://linkedlifedata.com/resource/pubmed/chemical/Organophosphorus Compounds, http://linkedlifedata.com/resource/pubmed/chemical/Phosphotransferases (Phosphate..., http://linkedlifedata.com/resource/pubmed/chemical/Polyphosphates, http://linkedlifedata.com/resource/pubmed/chemical/Transferases, http://linkedlifedata.com/resource/pubmed/chemical/deoxyxylulose-5-phosphate synthase, http://linkedlifedata.com/resource/pubmed/chemical/endopolyphosphatase, http://linkedlifedata.com/resource/pubmed/chemical/isopentenyl pyrophosphate, http://linkedlifedata.com/resource/pubmed/chemical/lycopene, http://linkedlifedata.com/resource/pubmed/chemical/polyphosphate kinase
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
1096-7176
pubmed:author
pubmed:issnType
Print
pubmed:volume
2
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
328-38
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
Low-copy plasmids can perform as well as or better than high-copy plasmids for metabolic engineering of bacteria.
pubmed:affiliation
Department of Chemical Engineering, University of California, Berkeley, California 94720-1462, USA.
pubmed:publicationType
Journal Article