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The article explores the use of the enzyme PestE from Pyrobaculum calidifontis VA1 for the enzymatic synthesis of hydroxycinnamic acid amides (HCAAs). HCAAs are natural compounds with potential applications in various industries. The current methods for synthesizing HCAAs are not environmentally friendly, so enzymatic methods are being explored as a greener alternative. The study focuses on the synthesis of N-trans-feruloyltyramine using PestE and found that the reaction was most efficient at 60 °C with a specific molar ratio. The use of acetonitrile as a cosolvent posed challenges, but ascorbic acid could be added to prevent substrate oxidation. PestE was also found to catalyze the formation of other hydroxycinnamic acid amides. Overall, this enzymatic method offers a more sustainable approach to HCAA synthesis. [Extracted from the article]

Promiskuitive Acyltransferase‐Aktivität ist die Eigenschaft mancher Hydrolasen, in wässriger Lösung Acyltransfer statt Hydrolyse zu katalysieren. Niedrige Enantioselektivität bzw. Effizienz, signifikante Produkthydrolyse und das begrenzte Substratspektrum bekannter Acyltransferasen stellen erhebliche Nachteile für industrielle Anwendungen dar. Durch aktivitätsbasiertes Screening von Hydrolasen konnte die Carboxylesterase EstCE1 aus der Familie VIII als beispiellos effiziente Acyltransferase identifiziert werden. EstCE1 kann die irreversible Amidierung und Carbamoylierung von Aminen in Wasser katalysieren, wie hier am Beispiel der Synthese des Wirkstoffs Moclobemid gezeigt. Detaillierte Struktur‐Funktions‐Analysen basierend auf der hier gelösten Kristallstruktur von EstCE1 identifizierten ein Sequenz‐Motiv, das sich als bedeutsam für die promiskuitive Acyltransferase‐Aktivität herausstellte. Die Übertragung dieses Motivs führte zur Umwandlung einer "Hydrolase" in eine "Acyltransferase". [ABSTRACT FROM AUTHOR]

Promiscuous acyltransferase activity is the ability of certain hydrolases to preferentially catalyze acyl transfer over hydrolysis, even in bulk water. However, poor enantioselectivity, low transfer efficiency, significant product hydrolysis, and limited substrate scope represent considerable drawbacks for their application. By activity‐based screening of several hydrolases, we identified the family VIII carboxylesterase, EstCE1, as an unprecedentedly efficient acyltransferase. EstCE1 catalyzes the irreversible amidation and carbamoylation of amines in water, which enabled the synthesis of the drug moclobemide from methyl 4‐chlorobenzoate and 4‐(2‐aminoethyl)morpholine (ca. 20 % conversion). We solved the crystal structure of EstCE1 and detailed structure–function analysis revealed a three‐amino acid motif important for promiscuous acyltransferase activity. Introducing this motif into an esterase without acetyltransferase activity transformed a "hydrolase" into an "acyltransferase". [ABSTRACT FROM AUTHOR]