Inverting enantioselectivity of Burkholderia gladioli esterase EstB by directed and designed evolution

Ivančić, Mirela; Valinger, Goran; Gruber, Karl; Schwab, Helmut

doi:10.1016/j.jbiotec.2006.10.007

Cited by 45 publications

(34 citation statements)

References 26 publications

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“…[55] Such pairwise mutagenesis in the active site may be the best strategy for reversing enantiopreference, as reversal involves cooperativity, and in this way the locations of two substituents of the chiral substrate can be changed. Indeed, multiple mutations have reversed the enantioselectivity of horseradish peroxidase, [56] the lipase from Burkholderia cepacia, [57,58] and esterases from Burkholderia gladioli [59] and Bacillus subtilis. [60] Of the nine examples in Table 2, only two involved a single mutation: A Phe352Val mutation in the [a] Raushel and co-workers also increased the S P selectivity of the wild-type enzyme to more than 100 with a Gly60Ala mutation, which decreased the size of the small subsite.…”

Section: Methodsmentioning

confidence: 99%

Enantiocomplementary Enzymes: Classification, Molecular Basis for Their Enantiopreference, and Prospects for Mirror‐Image Biotransformations

et al. 2008

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One often-cited weakness of biocatalysis is the lack of mirror-image enzymes for the formation of either enantiomer of a product in asymmetric synthesis. Enantiocomplementary enzymes exist as the solution to this problem in nature. These enzyme pairs, which catalyze the same reaction but favor opposite enantiomers, are not mirror-image molecules; however, they contain active sites that are functionally mirror images of one another. To create mirror-image active sites, nature can change the location of the binding site and/or the location of key catalytic groups. In this Minireview, X-ray crystal structures of enantiocomplementary enzymes are surveyed and classified into four groups according to how the mirror-image active sites are formed.

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Section: Methodsmentioning

confidence: 99%

Enantiocomplementary Enzymes: Classification, Molecular Basis for Their Enantiopreference, and Prospects for Mirror‐Image Biotransformations

et al. 2008

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“…[108] They demonstrated once again that a semirandom approach guided by the X-ray structure of the enzyme constitutes a powerful approach to fast laboratory evolution, in this case by first Figure 9. Binding pocket of IEMT reshaped by an ISM process.…”

Section: Inverting the Enantioselectivity Of The Esterase From Burkhomentioning

confidence: 99%

Laboratory Evolution of Stereoselective Enzymes: A Prolific Source of Catalysts for Asymmetric Reactions

2010

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Asymmetric catalysis plays a key role in modern synthetic organic chemistry, with synthetic catalysts and enzymes being the two available options. During the latter part of the last century the use of enzymes in organic chemistry and biotechnology experienced a period of rapid growth. However, these biocatalysts have traditionally suffered from several limitations, including in many cases limited substrate scope, poor enantioselectivity, insufficient stability, and sometimes product inhibition. During the last 15 years, the genetic technique of directed evolution has been developed to such an extent that all of these long-standing problems can be addressed and solved. It is based on repeated cycles of gene mutagenesis, expression, and screening (or selection). This Review focuses on the directed evolution of enantioselective enzymes, which constitutes a fundamentally new approach to asymmetric catalysis. Emphasis is placed on the development of methods to make laboratory evolution faster and more efficient, thus providing chemists and biotechnologists with a rich and non-ending source of robust and selective catalysts for a variety of useful applications.

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“…[108] Sie demonstrierten einmal mehr, dass die Kombination von Strukturdaten eines Enzyms und iterative Randomisierung an korrekt ausgewählten Positionen eine überzeugende Strategie in der gerichteten Evolution ist. Tatsächlich wurde zunächst epPCR getestet, aber dies war nicht sonderlich erfolgreich, um die (S)-Selektivität der WT-EstB in eine hoch (R)-selektive Mutante zu überführen.…”

Section: Umkehrung Der Enantioselektivität Der Esterase Aus Burkholdeunclassified

Gerichtete Evolution stereoselektiver Enzyme: Eine ergiebige Katalysator‐Quelle für asymmetrische Reaktionen

Reetz

2010

Angewandte Chemie

130

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Asymmetrische Katalyse spielt eine Schlüsselrolle in der modernen synthetischen organischen Chemie, wobei künstliche Katalysatoren und Enzyme die zwei wichtigsten Optionen darstellen. Die Verwendung von Enzymen in der organischen Chemie und Biotechnologie hat in der zweiten Hälfte des vergangenen Jahrhunderts stark zugenommen, oft blieb jedoch der Anwendungsbereich aus mehreren Gründen eingeschränkt. Dazu gehören begrenzte Substratakzeptanz, geringe oder fehlende Stereoselektivität, unzureichende Stabilität und manchmal Produktinhibierung. Während der vergangenen 15 Jahre wurde die genetische Technik der gerichteten Evolution so weit entwickelt und verfeinert, dass all diese Probleme angegangen und in der Regel gelöst werden können. Sie basiert auf wiederholende Zyklen von Genmutagenese, Expression und Screening (oder Selektion). Der vorliegende Aufsatz fokussiert auf gerichtete Evolution stereoselektiver Enzyme, ein fundamental neuer Ansatz zur asymmetrischen Katalyse. Der Schwerpunkt liegt auf Methodenentwicklung in dem Bestreben, diese Art der Katalysator‐Optimierung einfacher, schneller und effizienter als in der Vergangenheit zu gestalten. Die neueren Methoden bescheren dem Chemiker und dem Biotechnologen robuste und selektive Katalysatoren für eine Vielzahl von nützlichen Anwendungen.

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Inverting enantioselectivity of Burkholderia gladioli esterase EstB by directed and designed evolution

Cited by 45 publications

References 26 publications

Enantiocomplementary Enzymes: Classification, Molecular Basis for Their Enantiopreference, and Prospects for Mirror‐Image Biotransformations

Enantiocomplementary Enzymes: Classification, Molecular Basis for Their Enantiopreference, and Prospects for Mirror‐Image Biotransformations

Laboratory Evolution of Stereoselective Enzymes: A Prolific Source of Catalysts for Asymmetric Reactions

Gerichtete Evolution stereoselektiver Enzyme: Eine ergiebige Katalysator‐Quelle für asymmetrische Reaktionen

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