Isolation and properties of aromatic alpha-keto acid reductase.
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Animals↗
Biomedical subjects
W W Weber
Publications and source records attributed to W W Weber.
Reduction of phenylpyruvic acids to phenyllactic acids in mammalian tissues.
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Adrenal Glands↗
Linkage of Nat and Es-1 in the mouse and development of strains congenic for N-acetyltransferase.
The human polymorphism in the hepatic enzyme N-acetyltransferase (NAT) affects the rate at which individuals acetylate, and in many cases detoxify, aromatic amine and hydrazine drugs and xenobiotics. Differences in NAT activity are known to affect individual susceptibility to drug toxicities and are thought to play a part in some spontaneous disorders. A mouse model for the human acetylation polymorphism has been previously characterized and involves the A/J (slow acetylator) and C57BL/6J (rapid acetylator) inbred strains. Strain distribution analysis of 40 A x B and B x A recombinant inbred (RI) strains indicated linkage between the N-acetyltransferase gene (Nat) and the esterase 1 (Es-1) gene, located on mouse chromosome 8. A double backcross involving 107 animals confirmed the recombination frequency between Nat and Es-1 to be 12 +/- 3% (mean +/- SE). The information obtained in the backcross and RI studies was combined, yielding a 13 +/- 2.8% (mean +/- SD) recombination frequency. The Es-1 genotype was determined in our newly developed congenic strains A.B6-Natr and B6.A-Nats. The B6.A-Nats strain has the Es-1 genotype of its inbred partner, the B6 strain, and the A.B6-Natr strain has the Es-1 genotype of the donor strain. These congenic strains will be important in determining the role of the NAT genotype in susceptibility to arylamine-induced cancer and other disorders.
Acetyltransferases↗
Clinical pharmacokinetics of isoniazid.
The pharmacokinetics of isoniazid in man are described. Pronounced interindividual variation in circulating isoniazid concentration and clearance which occur after dosing with the drug are associated with hereditary differences in the acetylator status. The variations in rate of isoniazid inactivation and elimination in different (rapid and slow) acetylator phenotypes are primarily due to differences in the rate of acetylation of isoniazid by a genetically controlled polymorphic N-acetyltransferase in liver and small intestine. An appreciable 'first-pass' effect is observed following oral isoniazid administration, particularly in the rapid acetylator phenotype. Liver disease can cause a significant prolongation in the clearance of isoniazid; in the acutely ill patient, the prolongation correlates most closely with serum bilirubin elevation, although the degree of prolongation is less important than the intrinsic genetic difference between acetylator phenotypes. The effect of renal impairment on isoniazid excretion is relatively unimportant, even in slow acetylators. Methods for monitoring blood and urine concentrations of isoniazid and for acetylator phenotype determination which are convenient for the patient and clinician are available. Implications of phenotype differences in acetylator status for the optimal management of tuberculosis with isoniazid are considered. Attempts to devise new isoniazid formulations for this purpose are being evaluated.
Acetylation↗
Genetic polymorphism of cytochrome P-450-dependent phencyclidine hydroxylation in mice. Comparison of phencyclidine hydroxylation in humans.
A mouse model of P-450 phencyclidine 3-cyclohydroxylase (P-450 PCP 3-cyclohydroxylase) genetic polymorphism is described. Up to a 3-fold difference was observed in the constitutive liver microsomal activity of P-450 PCP 3-cyclohydroxylase between "slow" (A/J and DBA/2J) and "rapid" (C57BL/6J and BALB/CJ) phencyclidine (PCP) metabolizers. The segregation of slow and rapid hydroxylator phenotypes between 17 recombinant inbred mouse strains derived from A/J and C57BL/2J mice suggests control of the activity by a single gene located on the X-chromosome or, less likely, on chromosome 17. A liver deficiency of P-450 PCP 3-cyclohydroxylase was observed in the New Zealand rabbit and Wistar rat, as well as in some human subjects. Any relationship of P-450 PCP 3-cyclohydroxylase polymorphism to other well characterized P-450 polymorphisms in mice (aryl hydrocarbon hydroxylase and coumarin hydroxylase) was excluded on the basis of differences in inducibility and activity distribution among the inbred mouse strains. Lack of relationship to the P-450 debrisoquine hydroxylase was confirmed by direct comparison of both activities in the same mouse and human liver microsomes. The pharmacological consequence of the observed polymorphism in mice appears to be that the rapid PCP metabolizers are more resistant to the effects of PCP compared to the slow metabolizers as based upon its ED50 and duration of action in A/J and C57BL/6J mice. The relevance of this data to humans remains to be determined, but clearly the latter show marked differences in PCP 3-cyclohydroxylase activity, which separate into low, intermediate, and high groups.
Animals↗
Kinetics of acetyl CoA: arylamine N-acetyltransferase from rapid and slow acetylator human liver.
Michaelis-Menten kinetic constants for sulfamethazine (SMZ) and p-aminobenzoic acid (PABA) metabolism were determined in 2 very rapid, 8 intermediate, and 10 slow acetylator human livers. The mean apparent KM values for the monomorphic substrate PABA were 70 +/- 20, 180 +/- 50, and 310 +/- 30 microM for the slow, intermediate, and very rapid enzymes, respectively, whereas the polymorphic substrate SMZ exhibited little interphenotypic KM variation. Compared to the slow enzymes, the rapid acetylators exhibited mean apparent Vmax values 15- and 20-fold greater for PABA and SMZ, respectively. Furthermore, under in vitro conditions where enzyme saturation was achieved, PABA showed polymorphic acetylation characteristics, in contrast to events in vivo, where monomorphic acetylation patterns were observed. As the acetyl coenzyme A concentration in the reaction mixture was increased, the KM and Vmax for PABA increased, in accordance with "ping-pong" kinetic principles. As occurs with polymorphic substrates, a binary ping-pong mechanism appears to govern N-acetylation of monomorphic substrates in human liver, as evidenced by initial velocity patterns and limiting values for the KM and Vmax for PABA.
4-Aminobenzoic Acid↗
N-Acetylation of drugs. A genetically controlled reciprocal relationship between drug N-acetylating enzymes of rabbit liver and peripheral blood cells.
The reciprocal relation between liver isoniazid N-acetyltransferase and blood p-aminobenzoic and N-acetyltransferase previously reported is confirmed and found to be expressed in erythrocytes and lymphocytes of genetically rapid and slow isoniazid-acetylator rabbits. Both erythrocytes and lymphocytes from slow acetylator rabbits contained 2.5-3.3 times as much p-aminobenzoic acid N-acetyltransferase activity as the same cells from rapid acetylator rabbits. Mechanisms which might account for the reciprocal association between liver and blood N-acetyltransferases are considered.
4-Aminobenzoic Acid↗
Genetic differences in 2-aminofluorene pharmacokinetics between intact C57BL/6J and A/J mice.
Studies of genetically determined differences in arylamine acetylation with the model carcinogen 2-aminofluorene in C57BL/6J and A/J inbred mouse strains showed that individual differences in the pharmacokinetics of 2-aminofluorene were dependent on differences in 2-aminofluorene N-acetyltransferase activity in liver and blood. Elimination rates of 2-aminofluorene from blood of mice administered a single ip dose of 30, 50, or 100 mg/kg of 2-aminofluorene were dose-dependent in both strains. At a dose of 100 mg/kg, the average rate of 2-aminofluorene elimination was approximately three times faster in rapid acetylator (C57BL/6J) mice than in slow acetylator (A/J) mice (0.36 +/- 0.02 hr-1 vs. 0.12 +/- 0.02 hr-1), and that in B6AF1 mice was intermediate (0.24 +/- 0.02 hr-1) to the parental lines. These results support previous observations that acetylation of arylamines is controlled by intermediate dominant inheritance of two major alleles at a single locus. Comparison of the average rate of elimination of 2-aminofluorene from blood of the congenic mouse line, A.B6-NATr, (0.27 +/- 0.05 hr-1) which has the rapid acetylator allele placed on the A/J background provided evidence that modifying genes of the A/J mouse significantly reduced the effect of the rapid acetylator allele on the rate of 2-aminofluorene elimination. A trend was observed toward a greater apparent volume of distribution for 2-aminofluorene in A/J than in C57BL/6J mice.
Acetyltransferases↗
The molecular basis of hereditary acetylation polymorphisms.
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Acetylation↗
N-acetylation pharmacogenetics. Michaelis-Menten constants for arylamine drugs as predictors of their N-acetylation rates in vivo.
Michaelis-Menten constants for two in vivo monomorphically N-acetylated substrates, p-aminobenzoic acid and p-aminosalicylic acid, and two in vivo polymorphically N-acetylated substrates, sulfamethazine and procainamide, were determined with an improved assay procedure using liver N-acetyltransferase from rapid and slow acetylator rabbit. The slow rabbit liver isozyme proves to be a Vmax and a Km variant for p-aminobenzoic acid and p-aminosalicylic acid. Mean differences in the apparent Vmax for rapid acetylators were 39-fold greater for p-aminobenzoic acid and 16-fold greater for p-aminosalicylic acid. The apparent Km values for the slow acetylator enzyme were lower than 5 microM, whereas the apparent Km values for the rapid acetylator phenotype were at least 15 times higher, with a value of 105 +/- 21 microM for p-aminobenzoic acid and 74 +/- 16 microM for p-aminosalicylic acid. In contrast, for the polymorphic substrates, sulfamethazine and procainamide, rapid rabbit liver N-acetyltransferase was only a Vmax variant with a mean specific activity that was 13-fold higher than that for slow acetylator.
4-Aminobenzoic Acid↗
Genetic control of acetyl coenzyme A-dependent arylamine N-acetyltransferase, hydrazine N-acetyltransferase, and N-hydroxy-arylamine O-acetyltransferase enzymes in C57BL/6J, A/J, AC57F1, and the rapid and slow acetylator A.B6 and B6.A congenic inbred mouse.
Acetyl coenzyme A-dependent N-acetyltransferase and O-acetyltransferase activities were examined in liver cytosols derived from homozygous rapid acetylator C57BL/6J and A.B6 congenic inbred mouse strains, from homozygous slow acetylator A/J and B6.A congenic inbred mouse strains, and from the (C57BL/6J x A/J)F1 heterozygous acetylator hybrid mouse strain. Acetylator genotype-dependent N-acetyltransferase activity was exhibited for the N-acetylation of p-aminobenzoic acid, 2-aminofluorene, and 4-aminobiphenyl. In contrast, levels of isoniazid N-acetyltransferase and N-hydroxy-3,2'-dimethyl-4-aminobiphenyl O-acetyltransferase activities in mouse liver cytosol appeared to be independent of the arylamine Nat acetylator gene. Although cytosolic N-acetyltransferase activities differed about 2-fold between the parental C57BL/6J and A/J strains for p-aminobenzoic acid, 2-aminofluorene, and 4-aminobiphenyl, the same N-acetyltransferase activities differed about 6-7-fold between the homozygous rapid acetylator A.B6 and the homozygous slow acetylator B6.A congenic inbred strains. Partial purification of acetyl coenzyme A-dependent arylamine N-acetyltransferase activity in the five inbred mouse strains showed one major paraoxon-resistant enzyme in liver cytosol in each of the rapid and slow acetylator mouse strains examined. Levels of partially purified 2-aminofluorene and 4-aminobiphenyl N-acetyltransferase activity were about 7-fold higher in the A.B6 than the B6.A congenic inbred strain. Partial purification of acetyl coenzyme A-dependent isoniazid N-acetyltransferase activity showed catalysis by a paraoxon-resistant enzyme(s) distinct from the major arylamine N-acetyltransferase enzyme(s). These results suggest that isoniazid N-acetyltransferase(s) in mouse liver cytosol is a product of a separate gene that segregates independently of the arylamine Nat gene.(ABSTRACT TRUNCATED AT 250 WORDS)
Acetyl-CoA C-Acetyltransferase↗
N-acetylation of drugs. Pharmacogenetic studies in rabbits selected for their acetylator characteristics.
Studies on acetylation of sulfadiazine, isoniazid, and p-aminobenzoic acid in selected lines of slow and rapid acetylator rabbits are described. Pedigree analysis of rabbits classified as slow or rapid sulfadiazine acetylators confirmed previous studies that the rate of sulfadiazine elimination (acetylation) is genetically controlled, with rapid elimination dominant over slow elimination of the drug. Pharmacokinetic studies in rabbits of specified sulfadiazine acetylator genotypes with isoniazid and p-aminobenzoic acid show that the rate of isoniazid elimination is under the same genetic control as is sulfadiazine, whereas the rate of p-aminobenzoic acid elimination is not. A new drug acetylation polymorphism, which controls the rate of enzymatic acetylation of p-aminobenzoic acid in peripheral blood cells and which is related to the sulfadiazine acetylation polymorphism, is described.
Acetyltransferases↗
N-acetylation of drugs: search for INH acetylation polymorphism in baboons.
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Acetylation↗
Rodent models of the human isoniazid-acetylator polymorphism.
Inbred strains and subpopulations of rats, laboratory mice, and deer mice were examined for individual variation in the ability to metabolize several arylamines (p-aminobenzoic acid, sulfamethazine, aniline, alpha-naphthylamine, and aminofluorene) by N-acetylation. Individual differences within species were found to be dependent upon the tissue source of N-acetyltransferase activity and the acetyl acceptor employed. Long-Evans rats possessed about 2-fold more p-aminobenzoic acid N-acetyltransferase activity in blood and liver than Sprague-Dawley rats; no strain differences could be found with sulfamethazine. Nine strains of laboratory mice (Mus musculus) were found to have considerable liver p-aminobenzoic acid N-acetyltransferase activity but only slight activity towards sulfamethazine. No strain differences were apparent in regard to liver N-acetyltransferase activity. Blood p-aminobenzoic acid N-acetyltransferase activity was distinctly polymorphic in laboratory mice; of the nine strains tested, only A/J mice did not have this activity. Partially inbred deer mice (Peromyscus maniculatus) showed a narrower phenotypic range than random-bred stock from which they were obtained, which suggests the existence of distinct subpopulations with respect to N-acetylation capacity. Presumptive evidence for multiple forms of N-acetyltransferase in liver and blood was obtained through a study of substrate specificity.
Acetylation↗
Polymorphic N-acetylation of phenelzine and monoacetylhydrazine by highly purified rabbit liver isoniazid N-acetyltransferase.
Two hydrazines of clinical interest, the antidepressant drug phenelzine (PHZ) and the hepatotoxic metabolite of isoniazid, monoacetylhydrazine (MAH), were shown to be polymorphically acetylated by highly purified preparations of rabbit liver N-acetyltransferase (NAT). Both PHZ and MAH NAT activities exhibited purification and heat inactivation characteristics indistinguishable from genetically polymorphic NAT. Lineweaver-Burk analyses of PHZ and MAH NAT activities yielded apparent KM values of 4.18 mM for PHZ and 1.28 mM for MAH. These findings have implications concerning the structural requirements for polymorphic N-acetylation by liver NAT, and suggest that a wide variety of hydrazine compounds are also polymorphically N-acetylated.
Acetylation↗
N-acetylation polymorphism in liver and pancreas of inbred rats.
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Acetylation↗
Distribution of 2-aminofluorene and p-aminobenzoic acid N-acetyltransferase activity in tissues of C57BL/6J rapid and B6.A-NatS slow acetylator congenic mice.
The distribution of N-acetyltransferase (NAT) activity in 35 tissues of inbred rapid acetylator C57BL/6J and slow acetylator congenic B6.A-NatS mice was determined by incubation of tissue cytosols with 2-aminofluorene or p-aminobenzoic acid followed by HPLC assay. Tissues examined included the gastrointestinal tract, lymphoid tissues, skin, blood components, and other major organs. NAT activity was found in all tissues examined except blood plasma and seminal vesicles. Peyer's patches had the highest activity with either substrate, and lymphoid tissue, in general, was high in NAT activity as was skin and much of the digestive system. The acetylator polymorphism was apparent in most tissues for both p-aminobenzoic acid and 2-aminofluorene. The difference between rapid and slow acetylator phenotypes was usually greater with p-aminobenzoic acid than with 2-aminofluorene. The presence of NAT in the 33 tissues of rapid and slow acetylator mice, as well as the absence of NAT in plasma and seminal vesicles, was confirmed by immunoblots using an anti-NAT antibody raised in rabbits. These results indicate the widespread distribution of NAT activity and the relative abundance of extrahepatic N-acetylation capacity in the mouse.
4-Aminobenzoic Acid↗
Inhibition of human red blood cell N-acetyltransferase.
Acetanilide and derivatives of benzoic acid, benzamide, salicylic acid, and salicylamide were tested as inhibitors of p-aminobenzoic acid N-acetyltransferase partially purified from peripheral blood of three rapid and two slow dapsone acetylator human volunteers. The 50% inhibition point of these compounds does not provide a kinetic basis for differentiating the rapid and slow dapsone acetylator phenotypes.
4-Aminobenzoic Acid↗