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Amines to Amides: Acylation of Amines01:19

Amines to Amides: Acylation of Amines

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Various carboxylic acid derivatives (such as acid chlorides, esters, and anhydrides) can be used for the acylation of amines to yield amides. The reaction requires two equivalents of amines. The first amine molecule functions as a nucleophile and attacks the carbonyl carbon to produce a tetrahedral intermediate. This is followed by the loss of the leaving group and restoration of the C=O bond.
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Amides to Carboxylic Acids: Hydrolysis01:28

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Amides can undergo either acid-catalyzed hydrolysis or base-promoted hydrolysis through a typical nucleophilic acyl substitution. Each hydrolysis requires severe conditions.
Acid-catalyzed hydrolysis:
Hydrolysis of amides under acidic conditions yields carboxylic acids. Since the reaction occurs slowly, hydrolysis requires the conditions of heat.
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Lipids also are sources of energy that power cellular processes. Like carbohydrates, lipids are composed of carbon, hydrogen, and oxygen, but these atoms are arranged differently. Most lipids are nonpolar and hydrophobic. Major types include fats and oils, waxes, phospholipids, and steroids.
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Phase II Reactions: Acetylation Reactions01:24

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Acetylation, a phase II biotransformation reaction, introduces an acetyl group to drugs or their metabolites. Acetyltransferase enzymes facilitate this reaction, which resembles α-amino acid conjugation due to the addition of a functional group to the drug molecule.
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Preparation of Amides01:29

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Amides are synthesized by treating carboxylic acids with amines in the presence of dehydrating agents like dicyclohexylcarbodiimide (DCC).
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Acid Halides to Amides: Aminolysis01:07

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Aminolysis is a nucleophilic acyl substitution reaction, where ammonia or amines act as nucleophiles to give the substitution product. Acid halides react with ammonia, primary amines, and secondary amines to yield primary, secondary, and tertiary amides, respectively.
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Revisiting D-Acylases for D-Amino Acid Production.

Sergio Martínez-Rodríguez1,2, Jose Antonio Gavira2,3

  • 1Department of Biochemistry and Molecular Biology III and Immunology, University of Granada, Granada, Spain.

Microbial Biotechnology
|June 10, 2025
PubMed
Summary
This summary is machine-generated.

This study characterizes new D-acylases for dynamic kinetic resolution (DKR) of N-acyl-D,L-amino acids, enabling efficient D-amino acid biosynthesis. Structural analysis reveals a mobile domain crucial for enzyme specificity and protein engineering.

Keywords:
N‐acyl‐D‐amino acid deacylaseN‐acyl‐amino acidN‐succinyl‐amino acid racemaseacylaseamidohydrolaseamidohydrolase processamino acidenzyme cascaderacemase

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Area of Science:

  • Biocatalysis and Enzyme Engineering
  • Structural Biology
  • Biotechnology

Background:

  • N-Acyl-D-amino acid deacylases (D-acylases) are key enzymes for stereospecific resolution of N-acetyl-D,L-amino acids (NAAs).
  • Coupling D-acylases with N-succinyl-amino acid racemases (NSARs) facilitates dynamic kinetic resolution (DKR) for enantiomerically pure D-amino acid production.
  • Previous applications of D-acylase/NSAR systems have been primarily industrial, with limited exploration in broader research.

Purpose of the Study:

  • To characterize novel recombinant D-acylases from Bordetella petrii and Klebsiella pneumoniae.
  • To evaluate the efficacy of these D-acylases coupled with NSAR for D-amino acid biosynthesis.
  • To elucidate the structure of Klebsiella pneumoniae D-acylase (KleDacyl) and understand its catalytic mechanism and substrate specificity.

Main Methods:

  • Recombinant expression and characterization of two new D-acylases.
  • Enzymatic coupling of D-acylases with a recombinant NSAR from Geobacillus stearothermophilus.
  • Biosynthesis of D-methionine and D-aminobutyric acid.
  • X-ray crystallography for structural determination of KleDacyl.

Main Results:

  • Two novel D-acylases were successfully characterized and coupled with NSAR for D-amino acid production.
  • The study reports the second experimental 3-D structure of a D-acylase, KleDacyl, revealing a highly dynamic enzyme.
  • A mobile α/β domain (residues 282-341) was identified as critical for KleDacyl substrate specificity.

Conclusions:

  • D-acylase/NSAR tandems are effective biocatalytic tools for D-amino acid synthesis beyond industrial settings.
  • The determined structure of KleDacyl provides insights into enzyme dynamics and substrate binding mechanisms.
  • The identified mobile domain offers a target for protein engineering to tailor D-acylase specificity for novel applications.