Preparation of a Soluble Bifunctional Enzyme by Gene Fusion

Bülow, Leif; Ljungcrantz, Peter; Mosbach, Klaus

doi:10.1038/nbt0985-821

Cited by 59 publications

(33 citation statements)

References 18 publications

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“…In biocatalysis, one extensively studied cytochrome P450 monooxygenase is the natural fusion P450 BM3 from Bacillus megaterium . It can catalyze enantio‐ and regioselective hydroxylations, and it consists of two catalytic domains: the CPR, which is a NADPH‐dependent FAD‐ and FMN‐containing reductase domain, and a monooxygenase heme domain.…”

Section: Enzyme Fusions In Biocatalysismentioning

confidence: 58%

“…One of the first papers describes a fusion of a histidinol dehydrogenase with an aminotransferase . Another important early study was done on fusing β‐galactosidase with a galactose dehydrogenase; the fused enzyme showed improvement in the two‐step conversion from lactose to galactone . From this point on, many more enzyme fusions were produced and studied, of which some have been reviewed before .…”

Section: Enzyme Fusions In Biocatalysismentioning

confidence: 99%

“…[21] Another importante arly study wasd one on fusing b-galactosidase with ag alactose dehydrogenase;t he fused enzymes howedi mprovement in the two-stepc onversion from lactoset og alactone. [22] From this point on, many more enzyme fusions were produced and studied,o fw hich some have been reviewed before. [16,23] This Minireviewh ighlights some recent developments in fusing enzymes for biocatalytic applications.…”

Section: Enzyme Fusions In Biocatalysismentioning

confidence: 99%

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Enzyme Fusions in Biocatalysis: Coupling Reactions by Pairing Enzymes

2018

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One approach to bringing enzymes together for multienzyme biocatalysis is genetic fusion. This enables the production of multifunctional enzymes that can be used for whole-cell biotransformations or for in vitro (cascade) reactions. In some cases and in some aspects, such as expression and conversions, the fused enzymes outperform a combination of the individual enzymes. In contrast, some enzyme fusions are greatly compromised in activity and/or expression. In this Minireview, we give an overview of studies on fusions between two or more enzymes that were used for biocatalytic applications, with a focus on oxidative enzymes. Typically, the enzymes are paired to facilitate cofactor recycling or cosubstrate supply. In addition, different linker designs are briefly discussed. Although enzyme fusion is a promising tool for some biocatalytic applications, future studies could benefit from integrating the findings of previous studies in order to improve reliability and effectiveness.

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Section: Enzyme Fusions In Biocatalysismentioning

confidence: 58%

Section: Enzyme Fusions In Biocatalysismentioning

confidence: 99%

Section: Enzyme Fusions In Biocatalysismentioning

confidence: 99%

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Enzyme Fusions in Biocatalysis: Coupling Reactions by Pairing Enzymes

2018

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“…For example, Bulow et al constructed a bifunctional enzyme containing cascade reactions mediated by E. coli β-galactosidase and galactokinase (Bulow et al, 1985). This synthetic fusion enzyme displayed kinetic advantages (1.5-2.4-fold) over free enzyme mixtures.…”

Section: Stable Enzymes As Standardized Building Blocksmentioning

confidence: 98%

“…The simplest way to obtain multi-function enzymes is the creation of chimeric proteins, in which two or more cascade enzymes are combined by a linker to form a multi-functional single polypeptide (Bulow et al, 1985;Conrado et al, 2008). For example, Bulow et al constructed a bifunctional enzyme containing cascade reactions mediated by E. coli β-galactosidase and galactokinase (Bulow et al, 1985).…”

Section: Stable Enzymes As Standardized Building Blocksmentioning

confidence: 99%

Production of biofuels and biochemicals by in vitro synthetic biosystems: Opportunities and challenges

Zhang

2015

Biotechnology Advances

162

101

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The largest obstacle to the cost-competitive production of low-value and high-impact biofuels and biochemicals (called biocommodities) is high production costs catalyzed by microbes due to their inherent weaknesses, such as low product yield, slow reaction rate, high separation cost, intolerance to toxic products, and so on. This predominant whole-cell platform suffers from a mismatch between the primary goal of living microbes - cell proliferation and the desired biomanufacturing goal - desired products (not cell mass most times). In vitro synthetic biosystems consist of numerous enzymes as building bricks, enzyme complexes as building modules, and/or (biomimetic) coenzymes, which are assembled into synthetic enzymatic pathways for implementing complicated bioreactions. They emerge as an alternative solution for accomplishing a desired biotransformation without concerns of cell proliferation, complicated cellular regulation, and side-product formation. In addition to the most important advantage - high product yield, in vitro synthetic biosystems feature several other biomanufacturing advantages, such as fast reaction rate, easy product separation, open process control, broad reaction condition, tolerance to toxic substrates or products, and so on. In this perspective review, the general design rules of in vitro synthetic pathways are presented with eight supporting examples: hydrogen, n-butanol, isobutanol, electricity, starch, lactate,1,3-propanediol, and poly-3-hydroxylbutyrate. Also, a detailed economic analysis for enzymatic hydrogen production from carbohydrates is presented to illustrate some advantages of this system and the remaining challenges. Great market potentials will motivate worldwide efforts from multiple disciplines (i.e., chemistry, biology and engineering) to address the remaining obstacles pertaining to cost and stability of enzymes and coenzymes, standardized building parts and modules, biomimetic coenzymes, biosystem optimization, and scale-up, soon.

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Unique Applications of Immobilized Proteins in Bioanalytical Systems

Gupta¹,

Mattìasson²

1992

Methods of Biochemical Analysis

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Preparation of a Soluble Bifunctional Enzyme by Gene Fusion

Cited by 59 publications

References 18 publications

Enzyme Fusions in Biocatalysis: Coupling Reactions by Pairing Enzymes

Enzyme Fusions in Biocatalysis: Coupling Reactions by Pairing Enzymes

Production of biofuels and biochemicals by in vitro synthetic biosystems: Opportunities and challenges

Unique Applications of Immobilized Proteins in Bioanalytical Systems

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