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Allosteric Proteins-ATCase01:19

Allosteric Proteins-ATCase

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Binding sites linkages can regulate a protein's function.  For example, enzyme activity is often regulated through a feedback mechanism where the end product of the biochemical process serves as an inhibitor.
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to  N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis...
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Aminoacyl-tRNA synthetases are present in both eukaryotes and bacteria. Though eukaryotes have 20 different aminoacyl-tRNA synthetases to couple to 20 amino acids, many bacteria do not have genes for all of these aminoacyl-tRNA synthetases. Despite this, they still use all 20 amino acids to synthesize their proteins. For instance, some bacteria do not have the gene encoding the enzyme that couples glutamine with its partner tRNA. In these organisms, one enzyme adds glutamic acid to all of the...
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Ligand Binding and Linkage00:49

Ligand Binding and Linkage

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Allosteric proteins have more than one ligand binding site; the binding of a ligand to any of these sites influences the binding of ligands to the other sites. When a protein is allosteric, its binding sites are called coupled or linked.  In the case of enzymes, the site that binds to the substrate is known as the active site and the other site is known as the regulatory site. When a ligand binds to the regulatory site, this leads to conformational changes in the protein that can influence...
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Structure of Amines01:19

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The hybridized nitrogen atom in amines possesses a lone pair of electrons and is bound to three substituents with a bond angle of around 108°, which is less than the tetrahedral angle of 109.5°. However, the C–N–H bond angle is slightly larger at 112°, with a carbon–nitrogen bond length of 147 pm. This carbon–nitrogen bond length of of amines is longer than the carbon–oxygen bond of alcohols (143 pm) but shorter than alkanes’ carbon–carbon bond (154 pm). These aspects are...
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Ligand Binding Sites

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Proteins are dynamic macromolecules that carry out a wide variety of essential processes; however, the activities of most proteins depend on their interactions with other molecules or ions, known as ligands.
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Preparation of Amines: Alkylation of Ammonia and Amines

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Alkylation is one of the methods used to prepare amines. Direct alkylation of ammonia or a primary amine with an alkyl halide gives polyalkylated amines along with a quaternary ammonium salt through successive SN2 reactions. This process of making the quaternary salt through the direct alkylation method is called exhaustive alkylation.
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Efficient and Site-specific Antibody Labeling by Strain-promoted Azide-alkyne Cycloaddition
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Amine Transaminase Engineering for Spatially Bulky Substrate Acceptance.

Martin S Weiß1, Ioannis V Pavlidis1,2, Paul Spurr3

  • 1Department of Biotechnology and Enzyme Catalysis, Institute of Biochemistry, University of Greifswald, Felix-Hausdorff-Strasse 4, 17489, Greifswald, Germany.

Chembiochem : a European Journal of Chemical Biology
|March 24, 2017
PubMed
Summary

Engineered amine transaminases (ATAs) now efficiently convert bulky ketones into chiral amines. This breakthrough expands biocatalysis options beyond traditional methods for synthesizing valuable amine compounds.

Keywords:
amine transaminaseasymmetric synthesisenzyme catalysisprotein engineering

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Area of Science:

  • Biocatalysis
  • Organic Chemistry
  • Enzyme Engineering

Background:

  • Amine transaminases (ATAs) offer a sustainable alternative to transition metal catalysis for stereoselective amination.
  • Wild-type ATAs exhibit limited substrate scope, particularly with sterically hindered ketones.
  • Previous enzyme evolution efforts successfully broadened ATA substrate acceptance for planar bulky amines.

Purpose of the Study:

  • To develop engineered amine transaminases (ATAs) capable of accepting sterically demanding ketones, specifically those with tert-butyl substituents.
  • To overcome the inactivity of previously designed ATAs against substrates like 2,2-dimethyl-1-phenyl-propan-1-one.

Main Methods:

  • Protein engineering and directed evolution of amine transaminases.
  • Enzyme activity assays using challenging ketone substrates.
  • Asymmetric synthesis and product analysis.

Main Results:

  • Engineered ATAs demonstrated high conversion rates for the asymmetric synthesis of chiral amines from sterically hindered ketones.
  • The evolved enzymes successfully accepted 2,2-dimethyl-1-phenyl-propan-1-one, a previously challenging substrate.
  • High enantioselectivity was achieved for the synthesis of the (R)-amine enantiomer.

Conclusions:

  • The developed engineered ATAs provide a robust biocatalytic solution for the stereoselective synthesis of chiral amines from bulky ketones.
  • This advancement significantly expands the utility of amine transaminases in organic synthesis.
  • The findings pave the way for broader applications of biocatalysis in producing complex amine compounds.