ArticleThe Journal of biological chemistry2013
The mechanism of the amidases: mutating the glutamate adjacent to the catalytic triad inactivates the enzyme due to substrate mispositioning.
Article in The Journal of biological chemistry, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 16 papers.
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Who cites it
16 citing papers in PubMed, 45 citations in OpenAlex.
- Efficient acrylamide adduct formation suggests dual applications of ReAIV L-asparaginase.Amino acids · 2026Article
- Recombinant Amidases: Recent Insights and its Applications in the Production of Industrially Important Fine Chemicals.Molecular biotechnology · 2025Review
- Chemoselective, regioselective, and positionally selective fluorogenic stapling of unprotected peptides for cellular uptake and direct cell imaging.Chemical science · 2025Article
- Microbial amidases: Characterization, advances and biotechnological applications.Biotechnology notes (Amsterdam, Netherlands) · 2025Review
- The Allosteric Regulation of Β-Ureidopropionase Depends on Fine-Tuned Stability of Active-Site Loops and Subunit Interfaces.Biomolecules · 2023Article
- Molecular determinants of acrylamide neurotoxicity through covalent docking.Frontiers in pharmacology · 2023Article
- Effect of NaCl stress on exoproteome profiles ofFrontiers in microbiology · 2023Article
- Recent advances in the translation of drug metabolism and pharmacokinetics science for drug discovery and development.Acta pharmaceutica Sinica. B · 2022Review
- A Superefficient Ochratoxin A Hydrolase with Promising Potential for Industrial Applications.Applied and environmental microbiology · 2022Article
- Effective prediction of short hydrogen bonds in proteins via machine learning method.Scientific reports · 2022Article
- Different ways to transport ammonia in human and Mycobacterium tuberculosis NADNature communications · 2020Article
- Development and application of a transcriptional sensor for detection of heterologous acrylic acid production in E. coli.Microbial cell factories · 2019Article
- Cryo-EM and directed evolution reveal howCommunications biology · 2019Article
- Substrate specificity of plant nitrilase complexes is affected by their helical twist.Communications biology · 2018Article
- Crystal Structure of a Hidden Protein, YcaC, a Putative Cysteine Hydrolase from Pseudomonas aeruginosa, with and without an Acrylamide Adduct.International journal of molecular sciences · 2015Article
- Purification and characterization of a thermostable aliphatic amidase from the hyperthermophilic archaeon Pyrococcus yayanosii CH1.Extremophiles : life under extreme conditions · 2014Article
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Authors and funding
8 authors at 4 institutions in 3 countries.
Funding
Abstract
All known nitrilase superfamily amidase and carbamoylase structures have an additional glutamate that is hydrogen bonded to the catalytic lysine in addition to the Glu, Lys, Cys "catalytic triad." In the amidase from Geobacillus pallidus, mutating this glutamate (Glu-142) to a leucine or aspartate renders the enzyme inactive. X-ray crystal structure determination shows that the structural integrity of the enzyme is maintained despite the mutation with the catalytic cysteine (Cys-166), lysine (Lys-134), and glutamate (Glu-59) in positions similar to those of the wild-type enzyme. In the case of the E142L mutant, a chloride ion is located in the position occupied by Glu-142 O(ε1) in the wild-type enzyme and interacts with the active site lysine. In the case of the E142D mutant, this site is occupied by Asp-142 O(δ1.) In neither case is an atom located at the position of Glu-142 O(ε2) in the wild-type enzyme. The active site cysteine of the E142L mutant was found to form a Michael adduct with acrylamide, which is a substrate of the wild-type enzyme, due to an interaction that places the double bond of the acrylamide rather than the amide carbonyl carbon adjacent to the active site cysteine. Our results demonstrate that in the wild-type active site a crucial role is played by the hydrogen bond between Glu-142 O(ε2) and the substrate amino group in positioning the substrate with the correct stereoelectronic alignment to enable the nucleophilic attack on the carbonyl carbon by the catalytic cysteine.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.