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Transfer of the high-GC cyclohexane carboxylate degradation pathway from Rhodopseudomonas palustris to Escherichia coli for production of biotin

Paper ID Volume ID Publish Year Pages File Format Full-Text
31862 44846 2008 10 PDF Available
Title
Transfer of the high-GC cyclohexane carboxylate degradation pathway from Rhodopseudomonas palustris to Escherichia coli for production of biotin
Abstract

This work demonstrates the transfer of the five-gene cyclohexane carboxylate (CHC) degradation pathway from the high-GC alphaproteobacterium Rhodopseudomonas palustris to Escherichia coli, a gammaproteobacterium. The degradation product of this pathway is pimeloyl-CoA, a key metabolite in E. coli's biotin biosynthetic pathway. This pathway is useful for biotin overproduction in E. coli; however, the expression of GC-rich genes is troublesome in this host. When the native R. palustris CHC degradation pathway is transferred to a ΔbioH pimeloyl-CoA auxotroph of E. coli, it is unable to complement growth in the presence of CHC. To overcome this expression problem we redesigned the operon with decreased GC content and removed stretches of high-GC intergenic DNA which comprise the 5′ untranslated region of each gene, replacing these features with shorter low-GC sequences. We show this synthetic construct enables growth of the ΔbioH strain in the presence of CHC. When the synthetic degradation pathway is overexpressed in conjunction with the downstream genes for biotin biosynthesis, we measured significant accumulation of biotin in the growth medium, showing that the pathway transfer is successfully integrated with the host metabolism.

Keywords
Biotin; Synthetic DNA; Rhodopseudomonas palustris; Escherichia coli; High-GC; 5′ untranslated region (UTR)CHC, cyclohexane carboxylate; RBS, ribosome-binding sites; UTR, untranslated region; IPTG, isopropyl-β-d-thiogalactopyranoside; aTc, anhydrotetracycl
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Transfer of the high-GC cyclohexane carboxylate degradation pathway from Rhodopseudomonas palustris to Escherichia coli for production of biotin
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Publisher
Database: Elsevier - ScienceDirect
Journal: Metabolic Engineering - Volume 10, Issues 3–4, May–July 2008, Pages 131–140
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us