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Polymorphisms of N-acetyltransferase genes
Abstract:
1. A genetic polymorphism of human liver arylamine N-acetyltransferase (NAT) enzyme activity leads to wide variation in the disposition of many drugs and potential carcinogens, resulting in differential susceptibility to chemical-induced toxicity. 2. During studies to determine the biochemical and molecular mechanisms underlying this pharmacogenetic defect, we cloned two human genes, NAT1 and NAT2, which encode the functional acetylating enzymes NAT1 and NAT2. 3. NAT1 and NAT2 are both expressed in human liver cytosol, the latter as two closely related isoforms NAT2A and NAT2B. 4. NAT2 gene locus is the site of the human acetylation polymorphism, because its products NAT2A and NAT2B selectively acetylate 'polymorphic' arylamine substrates (e.g. sulphamethazine), and since the liver content of these isozymes is markedly reduced in genetically slow acetylator subjects. 5. NAT1 shows marked kinetic selectivity for 'monomorphic' substrates (e.g. p-aminobenzoic acid) whose in vivo acetylation rates do not correlate with the acetylation polymorphism. 6. Despite the drastic reduction in NAT2A/B proteins in livers from phenotypically slow acetylators, levels of the NAT2 gene transcript are not altered. 7. Three common mutant alleles at the NAT2 gene locus have so far been identified, which may be detected by restriction fragment length polymorphism (RFLP) analysis on Southern blots or by allele-specific polymerase chain reaction amplification.
Insights
Human liver enzyme activity, arylamine N-acetyltransferase (NAT), varies due to genetic polymorphism, affecting drug response and toxicity. The NAT2 gene locus is identified as the primary site of this human acetylation polymorphism.
Area of Science:
- Pharmacogenetics
- Biochemistry
- Molecular Biology
Background:
- Genetic variations in human liver arylamine N-acetyltransferase (NAT) enzyme activity cause significant differences in drug metabolism and carcinogen disposition.
- This pharmacogenetic variation leads to differential susceptibility to chemically induced toxicity.
- Understanding these mechanisms is crucial for personalized medicine and risk assessment.
Purpose of the Study:
- To elucidate the biochemical and molecular mechanisms underlying the human acetylation polymorphism.
- To identify and characterize the genes responsible for NAT enzyme activity.
- To investigate the genetic basis of differential drug and carcinogen metabolism.
Main Methods:
- Cloning of human NAT1 and NAT2 genes.
- Analysis of NAT1 and NAT2 expression in human liver cytosol.
- Characterization of NAT2 isoforms (NAT2A and NAT2B) and their substrate specificity.
- Assessment of liver enzyme content in slow vs. rapid acetylator phenotypes.
- Analysis of NAT2 gene transcript levels in slow acetylators.
- Identification and detection of mutant NAT2 alleles using RFLP and allele-specific PCR.
Main Results:
- Two human genes, NAT1 and NAT2, encoding functional acetylating enzymes NAT1 and NAT2, were cloned.
- NAT1 and NAT2 are expressed in human liver cytosol, with NAT2 existing as isoforms NAT2A and NAT2B.
- The NAT2 gene locus is responsible for the human acetylation polymorphism, as its products selectively acetylate polymorphic substrates and are reduced in slow acetylators.
- NAT1 exhibits kinetic selectivity for monomorphic substrates, independent of the acetylation polymorphism.
- Reduced NAT2A/B protein levels in slow acetylators are not associated with altered NAT2 gene transcript levels.
- Three common mutant NAT2 alleles have been identified.
Conclusions:
- The NAT2 gene locus is the primary determinant of the human acetylation polymorphism.
- Phenotypic slow acetylators exhibit reduced levels of NAT2A/B proteins, despite normal NAT2 transcript levels.
- Mutations within the NAT2 gene locus underlie the observed acetylation polymorphism, offering potential for genotypic detection and prediction of metabolic status.