Regulatory effects of cellular nicotinamide nucleotides and enzyme activities on poly(3-hydroxybutyrate) synthesis in recombinant Escherichia coli

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Title
Regulatory effects of cellular nicotinamide nucleotides and enzyme activities on poly(3-hydroxybutyrate) synthesis in recombinant Escherichia coli
Author(s)
In Young Lee; Mi Kyung Kim; Young Hoon Park; Sang Yup Lee
Bibliographic Citation
Biotechnology and Bioengineering, vol. 52, pp. 707-712
Publication Year
1996
Abstract
Regulatory roles of nicotinamide nucleotides and three key enzymes, β- ketothiolase (KT), NADPH-dependent acetoacetyl-CoA reductase (AAR), and citrate synthase (CS), on poly(3-hydroxybutyrate) (PHB) synthesis in recombinant Escherichia coli harboring a plasmid containing the Alcaligenes eutrophus polyhydroxyalkanoate (PHA) biosynthesis genes were examined. Cells were grown in various media and were subsequently compared for PHB concentration, PHB content, the activities of the key enzymes, and the levels of nicotinamide nucleotides. Cells of recombinant E. coli accumulated the largest amount of PHB in LB+glucose medium among those tested. PHB synthesis was not enhanced by limiting inorganic ions. The activity of CS, which competes with KT for acetyl-CoA, was lower when cells were grown in LB+glucose compared with other media. The NADPH level and the NADPH/NADP ratio were high in LB+glucose. Examination of the time profiles of the specific PHB synthesis rate, key enzyme activities, and the levels of nicotinamide nucleotides showed that PHB synthesis is most significantly affected by the NADPH level. Even though the NADH level and the NADH/NAD ratio were also high during the synthesis of PHB, no direct evidence of their positive effect on PHB synthesis was found. Low activity of CS was beneficial for PHB synthesis due to the availability of more acetyl-CoA to PHB biosynthetic pathway. In recombinant E. coli, the level of NADPH and/or the NADPH/NADP ratio seem to be the most critical factor regulating the activity of AAR and, subsequently, PHB synthesis.
Keyword
poly(3-hydroxybutyrate)regulationescherichia colinicotinamide nucleotidesPHB synthesis
ISSN
0006-3592
Publisher
Wiley
DOI
http://dx.doi.org/10.1002/(SICI)1097-0290(19961220)52:6<707::AID-BIT8>3.0.CO;2-S
Type
Article
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1. Journal Articles > Journal Articles
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