From random mutagenesis to systems biology in metabolic engineering of mammalian cells

2014 ◽  
Vol 2 (5) ◽  
pp. 355-358 ◽  
Author(s):  
Hooman Hefzi ◽  
Nathan E Lewis
Keyword(s):  
Metabolic Engineering ◽  
Systems Biology ◽  
Mammalian Cells ◽  
Random Mutagenesis

scholarly journals Random Mutagenesis of Clostridium cellulolyticum by Using a Tn1545 Derivative

2010 ◽  
Vol 76 (13) ◽  
pp. 4546-4549 ◽  
Author(s):  
Jean-Charles Blouzard ◽  
Odile Valette ◽  
Chantal Tardif ◽  
Pascale de Philip
Keyword(s):  
Cell Wall ◽  
Metabolic Engineering ◽  
Plant Cell ◽  
Plant Cell Wall ◽  
Random Mutagenesis ◽  
Cell Wall Degradation ◽  
Clostridium Cellulolyticum ◽  
Plant Cell Wall Degradation ◽  
Random Insertion

ABSTRACT Further understanding of the plant cell wall degradation system of Clostridium cellulolyticum and the possibility of metabolic engineering in this species highlight the need for a means of random mutagenesis. Here, we report the construction of a Tn1545-derived delivery tool which allows monocopy random insertion within the genome.

Systems biology of mammalian cells: A report from the Freiburg conference

BioEssays ◽  
2010 ◽  
Vol 32 (12) ◽  
pp. 1099-1104 ◽  
Author(s):  
Jens Timmer ◽  
Adriano Henney ◽  
Andrew Moore ◽  
Ursula Klingmuller
Keyword(s):  
Systems Biology ◽  
Mammalian Cells

scholarly journals Systems biology of virus entry in mammalian cells

2006 ◽  
Vol 8 (8) ◽  
pp. 1219-1227 ◽  
Author(s):  
Eva-Maria Damm ◽  
Lucas Pelkmans
Keyword(s):  
Systems Biology ◽  
Mammalian Cells ◽  
Virus Entry

The usefulness of post-genomics tools for characterization of the amine cross-talk in mammalian cells

2007 ◽  
Vol 35 (2) ◽  
pp. 381-385 ◽  
Author(s):  
F. Sánchez-Jiménez ◽  
R. Montañez ◽  
F. Correa-Fiz ◽  
P. Chaves ◽  
C. Rodríguez-Caso ◽  
...  
Keyword(s):  
Systems Biology ◽  
Mathematical Models ◽  
Mammalian Cells ◽  
State Of The Art ◽  
Molecular Approaches ◽  
Amine Metabolism ◽  
Genomics Tools ◽  
Stored Information

Evidence is growing in favour of a relationship between cancer and chronic inflammation, and particularly of the role of a polyamine and histamine metabolic interplay involved in these physiopathological problems, which are indeed highly complex biological systems. Decodification of the complex inter- and intra-cellular signalling mechanisms that control these effects is not an easy task, which must be helped by systems biology technologies, including new tools for location and integration of database-stored information and predictive mathematical models, as well as functional genomics and other experimental molecular approaches necessary for hypothesis validation. We review the state of the art and present our latest efforts in this area, focused on the amine metabolism field.

scholarly journals A colourless green fluorescent protein homologue from the non-fluorescent hydromedusa Aequorea coerulescens and its fluorescent mutants

2003 ◽  
Vol 373 (2) ◽  
pp. 403-408 ◽  
Author(s):  
Nadya G. GURSKAYA ◽  
Arkady F. FRADKOV ◽  
Natalia I. POUNKOVA ◽  
Dmitry B. STAROVEROV ◽  
Maria E. BULINA ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Mammalian Cells ◽  
Fluorescent Protein ◽  
Random Mutagenesis ◽  
Wild Type ◽  
Green Luminescence ◽  
Mammalian Cell Lines ◽  
Biological Tool ◽  
Fluorescent Pattern ◽  
Green Fluorescent

We have cloned an unusual colourless green fluorescent protein (GFP)-like protein from Aequorea coerulescens (acGFPL). The A. coerulescens specimens displayed blue (not green) luminescence, and no fluorescence was detected in these medusae. Escherichia coli expressing wild-type acGFPL showed neither fluorescence nor visible coloration. Random mutagenesis generated green fluorescent mutants of acGFPL, with the strongest emitters found to contain an Glu222→Gly (E222G) substitution, which removed the evolutionarily invariant Glu222. Re-introduction of Glu222 into the most fluorescent random mutant, named aceGFP, converted it into a colourless protein. This colourless aceGFP-G222E protein demonstrated a novel type of UV-induced photoconversion, from an immature non-fluorescent form into a green fluorescent form. Fluorescent aceGFP may be a useful biological tool, as it was able to be expressed in a number of mammalian cell lines. Furthermore, expression of a fusion protein of ‘humanized’ aceGFP and β-actin produced a fluorescent pattern consistent with actin distribution in mammalian cells.

scholarly journals Application of Random Mutagenesis and Synthetic FadR Promoter for de novo Production of ω-Hydroxy Fatty Acid in Yarrowia lipolytica

2021 ◽  
Vol 9 ◽  
Author(s):  
Beom Gi Park ◽  
Junyeob Kim ◽  
Eun-Jung Kim ◽  
Yechan Kim ◽  
Joonwon Kim ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Metabolic Engineering ◽  
Palmitic Acid ◽  
Yarrowia Lipolytica ◽  
Cell Sorting ◽  
De Novo ◽  
Random Mutagenesis ◽  
Oleaginous Yeasts ◽  
Synthetic Promoters ◽  
Base Strain

As a means to develop oleaginous biorefinery, Yarrowia lipolytica was utilized to produce ω-hydroxy palmitic acid from glucose using evolutionary metabolic engineering and synthetic FadR promoters for cytochrome P450 (CYP) expression. First, a base strain was constructed to produce free fatty acids (FFAs) from glucose using metabolic engineering strategies. Subsequently, through ethyl methanesulfonate (EMS)-induced random mutagenesis and fluorescence-activated cell sorting (FACS) screening, improved FFA overproducers were screened. Additionally, synthetic promoters containing bacterial FadR binding sequences for CYP expression were designed to respond to the surge of the concentration of FFAs to activate the ω-hydroxylating pathway, resulting in increased transcriptional activity by 14 times from the third day of culture compared to the first day. Then, endogenous alk5 was screened and expressed using the synthetic FadR promoter in the developed strain for the production of ω-hydroxy palmitic acid. By implementing the synthetic FadR promoter, cell growth and production phases could be efficiently decoupled. Finally, in batch fermentation, we demonstrated de novo production of 160 mg/L of ω-hydroxy palmitic acid using FmeN3-TR1-alk5 in nitrogen-limited media. This study presents an excellent example of the production of ω-hydroxy fatty acids using synthetic promoters with bacterial transcriptional regulator (i.e., FadR) binding sequences in oleaginous yeasts.

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