Biotransformation of crotonobetaine to L(-)- carnitine in Proteus sp.

Engemann, Claudia; Elßner, Thomas; Kleber, Hans‐Peter

doi:10.1007/s002030100272

Cited by 12 publications

(13 citation statements)

References 17 publications

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“…The largest differences between the enzymes relate to their quaternary structures, which vary considerably. The known enzymes of the family are either monomers (oxalate CoA‐transferase [18]), homodimers (CaiB [26–28]) or form more complex structures, such as the α 2 β 2 structure of benzylsuccinate CoA‐transferase [17] or the enzyme complex of phenyllactate CoA‐transferase and phenyllactyl‐CoA dehydratase [16]. One of the related proteins of unknown function from M. tuberculosis (accession number CAB06121) even seems to be a fusion of two subdomains that are both similar to CoA‐transferases of the new family.…”

Section: Discussionmentioning

confidence: 99%

A new family of CoA‐transferases

2001

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CoA-transferases are found in organisms from all lines of descent. Most of these enzymes belong to two well-known enzyme families, but recent work on unusual biochemical pathways of anaerobic bacteria has revealed the existence of a third family of CoA-transferases. The members of this enzyme family differ in sequence and reaction mechanism from CoAtransferases of the other families. Currently known enzymes of the new family are a formyl-CoA : oxalate CoA-transferase, a succinyl-CoA : (R)-benzylsuccinate CoA-transferase, an (E)-cinnamoyl-CoA : (R)-phenyllactate CoA-transferase, and a butyrobetainyl-CoA: (R)-carnitine CoA-transferase. In addition, a large number of proteins of unknown or differently annotated function from Bacteria, Archaea and Eukarya apparently belong to this enzyme family. Properties and reaction mechanisms of the CoA-transferases of family III are described and compared to those of the previously known CoA-transferases. ß

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Section: Discussionmentioning

confidence: 99%

A new family of CoA‐transferases

2001

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“…[90]. This transformation, along with its analogue in E. coli [91], is, however, reversible and constitutes a pathway of carnitine degradation in microbes.…”

Section: Production Of Optically Active Compoundsmentioning

confidence: 99%

Biotransformations Utilizing β-Oxidation Cycle Reactions in the Synthesis of Natural Compounds and Medicines

Œwizdor

Panek

Milecka-Tronina

et al. 2012

IJMS

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β-Oxidation cycle reactions, which are key stages in the metabolism of fatty acids in eucaryotic cells and in processes with a significant role in the degradation of acids used by microbes as a carbon source, have also found application in biotransformations. One of the major advantages of biotransformations based on the β-oxidation cycle is the possibility to transform a substrate in a series of reactions catalyzed by a number of enzymes. It allows the use of sterols as a substrate base in the production of natural steroid compounds and their analogues. This route also leads to biologically active compounds of therapeutic significance. Transformations of natural substrates via β-oxidation are the core part of the synthetic routes of natural flavors used as food additives. Stereoselectivity of the enzymes catalyzing the stages of dehydrogenation and addition of a water molecule to the double bond also finds application in the synthesis of chiral biologically active compounds, including medicines. Recent advances in genetic, metabolic engineering, methods for the enhancement of bioprocess productivity and the selectivity of target reactions are also described.

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“…The natural enantioselectivity of microbial and enzymatic biotransformations offers an advantage over classical chemical synthesis. Several biological processes have been developed for the production of L(-)-carnitine from non-chiral precursors [5-12] especially using strains belonging to the genera Escherichia and Proteus. At the industrial level , Lonza belongs a proprietary strain of a non-disclosed genus branching between Agrobacterium and Rhizobium and close to Rhizobium meliloti [13] .…”

Section: Introductionmentioning

confidence: 99%

Metabolic engineering for high yielding L(-)-carnitine production in Escherichia coli

et al. 2013

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BackgroundL(-)-carnitine production has been widely studied because of its beneficial properties on various diseases and dysfunctions. Enterobacteria possess a specific biotransformation pathway which can be used for the enantioselective production of L(-)-carnitine. Although bioprocesses catalyzed by enzymes or whole cells can overcome the lack of enantioselectivity of chemical methods, current processes for L(−)-carnitine production still have severe disadvantages, such as the low yields, side reactions and the need of high catalyst concentrations and anaerobic conditions for proper expression of the biotransformation pathway. Additionally, genetically engineered strains so far constructed for L(-)-carnitine production are based on plasmids and, therefore, suffer from segregational unstability.ResultsIn this work, a stable, high yielding strain for L(-)-carnitine production from low cost substrates was constructed. A metabolic engineering strategy was implemented in a multiple mutant for use in both growing and resting cells systems. The effect of mutations on gene expression and metabolism was analyzed to characterize the productivity constraints of the wild type and the overproducer strains. Precise deletion of genes which encode proteins of central and carnitine metabolisms were performed. Specifically, flux through the TCA cycle was increased by deletion of aceK (which encodes a bifunctional kinase/phosphatase which inhibits isocitrate dehydrogenase activity) and the synthesis of the by-product γ-butyrobetaine was prevented by deletion of caiA (which encodes a crotonobetainyl-CoA reductase). Both mutations led to improve the L(-)-carnitine production by 20 and 42%, respectively. Moreover, the highly regulated promoter of the cai operon was substituted by a constitutive artificial promoter increasing the biotransformation rate, even under aerobic conditions. Resting cells of the BW ΔaceK ΔcaiA p37cai strain produced 59.6 mmol l-1 · h-1 of L(−)-carnitine, doubling the productivity of the wild type strain. In addition, almost total conversion was attained in less than two hours without concomitant production of the side product γ–butyrobetaine.ConclusionsL(-)-carnitine production has been enhanced by strain engineering. Metabolic engineering strategies herein implemented allowed obtaining a robust and high yielding E. coli strain. The new overproducer strain attained almost complete conversion of crotonobetaine into L(-)-carnitine with growing and resting cells, and even under aerobic conditions, overcoming the main environmental restriction to carnitine metabolism expression. So far, this is the best performing L(-)-carnitine production E. coli strain described.

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Biotransformation of crotonobetaine to L(-)- carnitine in Proteus sp.

Cited by 12 publications

References 17 publications

A new family of CoA‐transferases

A new family of CoA‐transferases

Biotransformations Utilizing β-Oxidation Cycle Reactions in the Synthesis of Natural Compounds and Medicines

Metabolic engineering for high yielding L(-)-carnitine production in Escherichia coli

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