31416-98-5Relevant articles and documents
Molecular identification of NAT8 as the enzyme that acetylates cysteine S-conjugates to mercapturic acids
Veiga-da-Cunha, Maria,Tyteca, Donatienne,Stroobant, Vincent,Courtoy, Pierre J.,Opperdoes, Fred R.,Van Schaftingen, Emile
experimental part, p. 18888 - 18898 (2011/04/17)
Our goal was to identify the reaction catalyzed by NAT8 (N-acetyltransferase 8), a putative N-acetyltransferase homologous to the enzyme (NAT8L) that produces N-acetylaspartate in brain. The almost exclusive expression of NAT8 in kidney and liver and its predicted association with the endoplasmic reticulum suggested that it was cysteinyl-S-conjugate N-acetyltransferase, the microsomal enzyme that catalyzes the last step of mercapturic acid formation. In agreement, HEK293T extracts of cells overexpressing NAT8 catalyzed the N-acetylation of S-benzyl-L-cysteine and leukotriene E4, two cysteine conjugates, but were inactive on other physiological amines or amino acids. Confocal microscopy indicated that NAT8 was associated with the endoplasmic reticulum. Neither of the two frequent single nucleotide polymorphisms found in NAT8, E104K nor F143S, changed the enzymatic activity or the expression of the protein by≥2-fold, whereas a mutation (R149K) replacing an extremely conserved arginine suppressed the activity. Sequencing of genomic DNA and EST clones corresponding to the NAT8B gene, which resulted from duplication of the NAT8 gene in the primate lineage, disclosed the systematic presence of a premature stop codon at codon 16. Furthermore, truncated NAT8B and NAT8 proteins starting from the following methionine (Met-25) showed no cysteinyl-S-conjugate N-acetyltransferase activity when transfected in HEK293T cells. Taken together, these findings indicate that NAT8 is involved in mercapturic acid formation and confirm that NAT8B is an inactive gene in humans. NAT8 homologues are found in all vertebrate genomes, where they are often encoded by multiple, tandemly repeated genes as many other genes encoding xenobiotic metabolism enzymes.
Leaving Group Effects in Thiolester Alkaline Hydrolysis. Part 1. A Keten-mediated (E1cB) Pathway for Basic Hydrolysis of S-Acetoacetylcoenzyme A and Analogues
Douglas, Kenneth T.,Yaggi, Norbert F.
, p. 1037 - 1044 (2007/10/02)
The basic hydrolysis of a series of leaving-group substituted acetothiolacetates (CH3COCH2COSR) has been studied in aqueous media.Hydrolysis of N-acetyl-S-acetoacetylcysteinamine follows a kinetic ionisation curve with an inflexion corresponding to the pK of this ester as determined by spectrophotometric and electrometric titrations.The rate constant at high pH was shown to follow a Broensted relationship with βL.G. -1.13, where βL.G. is the slope of a plot of the logarithm of the rate constant versus the pKa of the conjugate acid of the leaving group.This, and other evidence from rate comparisons, activation parameters, and kinetic solvent isotope effects, indicated an E1cB hydrolytic mechanism involving unimolecular collapse of the ester enolate ions via a ketenoid transition-state.S-Acetoacetylcoenzyme-A was also hydrolysed in base by this mechanism.Direct comparison of rates of leaving group expulsion for ArS and ArO was possible by means of this unimolecular process.For a leaving group with pKL.G. 10, the oxyanion departs ca. 1 or 2 orders of magnitude faster than the thiolate anion; for pKL.G. 6.0, the advantage of oxygen over sulphur is 103-104 fold.In a direct structural comparison, PhS departs 32 times as rapidly as PhO.The contribution of steric release in the E1cB transition-state for S-t-butyl acetothiolacetate hydrolysis is discussed.The pKa values of some acetothiolacetates were measured.
