Slow acetylation in mice is caused by a labile and catalytically impaired mutant N-acetyltransferase (NAT2 9).
Three N-acetyltransferase genes (NAT*) were detected in inbred parental and congenic mice. Direct sequencing of NAT2* and liver cytosolic N-acetylation activity determinations with NAT2-specific (p-aminobenzoic acid) and NAT2-selective (2-aminofluorene) substrates have established that the acetylator congenic A.B6 and B6.A mice are genotypically and phenotypically identical to the parental B6 ("wild-type"; rapid acetylator) and A (mutant; slow acetylator) mice, respectively, from which they originated. The apparent KM for p-aminobenzoic acid and thermal inactivation rates determined with liver cytosol from the mutant (A and B6.A) mice were 3-fold and one order of magnitude higher than the corresponding values with liver cytosol from the wild-type (B6 and A.B6) strains. Northern blotting and immunoblotting revealed hepatic NAT2 mRNA and protein bands of equal size and intensity, regardless of the NAT2* genotype or phenotype of the animals. Incubation of liver cytosol from mutant A and B6.A mice at 37 degrees C for 6 hr resulted in virtual cessation of p-aminobenzoate N-acetylation activity, whereas the steady-state level of immunoreactive NAT2 remained unchanged. The results indicate that the amino acid change (N99I) in mutant NAT2* from slow acetylator mice does not hinder the synthesis of hepatic NAT2 protein, but, rather, leads to production of a conformationally modified NAT2 molecule that resists degradation by tissue proteases but is labile and catalytically impaired.
De Leon
J H
JH
Department of Pharmacology, University of Michigan.
Martell
K J
KJ
Vatsis
K P
KP
Weber
W W
WW
D000107:Acetylation; D000123:Acetyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D005260:Female; D005838:Genotype; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008297:Male; D051379:Mice; D008805:Mice, Inbred A; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D009154:Mutation / Q000502:physiology*; D010641:Phenotype; D010957:Plasmids; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis
D012333:RNA, Messenger; D000123:Acetyltransferases