Document Detail


High cell density cultivation of a recombinant E. coli strain expressing a key enzyme in bioengineered heparin production.
MedLine Citation:
PMID:  23318839     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
A bioengineered heparin, as a replacement for animal-derived heparin, is under development that relies on the fermentative production of heparosan by Escherichia coli K5 and its subsequent chemoenzymatic modification using biosynthetic enzymes. A critical enzyme in this pathway is the mammalian 6-O-sulfotransferase (6-OST-1) which specifically sulfonates the glucosamine residue in a heparin precursor. This mammalian enzyme, previously cloned and expressed in E. coli, is required in kilogram amounts if an industrial process for bioengineered heparin is to be established. In this study, high cell density cultivation techniques were exploited to obtain recombinant 6-OST-1. Physiological studies were performed in shake flasks to establish optimized growth and production conditions. Induction strategies were tested in fed-batch experiments to improve yield and productivity. High cell density cultivation in 7-l culture, together with a coupled inducer strategy using isopropyl β-D-1-thiogalactopyranoside and galactose, afforded 482 mg l(-1) of enzyme with a biomass yield of 16.2 mg gcdw (-1) and a productivity of 10.5 mg l(-1) h(-1).
Authors:
Odile Francesca Restaino; Ujjwal Bhaskar; Priscilla Paul; Lingyun Li; Mario De Rosa; Jonathan S Dordick; Robert J Linhardt
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't     Date:  2013-01-15
Journal Detail:
Title:  Applied microbiology and biotechnology     Volume:  97     ISSN:  1432-0614     ISO Abbreviation:  Appl. Microbiol. Biotechnol.     Publication Date:  2013 May 
Date Detail:
Created Date:  2013-04-16     Completed Date:  2013-10-21     Revised Date:  2014-05-07    
Medline Journal Info:
Nlm Unique ID:  8406612     Medline TA:  Appl Microbiol Biotechnol     Country:  Germany    
Other Details:
Languages:  eng     Pagination:  3893-900     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Arylsulfotransferase / genetics,  metabolism
Biomedical Engineering / methods*
Disaccharides / metabolism
Escherichia coli / enzymology*,  genetics,  metabolism*
Heparin / biosynthesis*
Grant Support
ID/Acronym/Agency:
HL096972/HL/NHLBI NIH HHS; R01 HL096972/HL/NHLBI NIH HHS
Chemical
Reg. No./Substance:
0/Disaccharides; 0/heparosan; 9005-49-6/Heparin; EC 2.8.2.1/Arylsulfotransferase
Comments/Corrections

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