New spectrophotometric and radiochemical assays for acetyl-CoA: arylamine N-acetyltransferase applicable to a variety of arylamines

Journal Article · Fri Mar 01 00:00:00 EST 1985 · Anal. Biochem.; (United States)

DOI:https://doi.org/10.1016/0003-2697(85)90376-8· OSTI ID:5253617

Andres, H H; Klem, A J; Szabo, S M; Weber, W W

Simple and sensitive spectrophotometric and radiochemical procedures are described for the assay of acetyl-CoA:arylamine N-acetyltransferase (NAT), which catalyzes the reaction acetyl-CoA + arylamine----N-acetylated arylamine + CoASH. The methods are applicable to crude tissue homogenates and blood lysates. The spectrophotometric assay is characterized by two features: (i) NAT activity is measured by quantifying the disappearance of the arylamine substrate as reflected by decreasing Schiff's base formation with dimethylaminobenzaldehyde. (ii) During the enzymatic reaction, the inhibitory product CoASH is recycled by the system acetyl phosphate/phosphotransacetylase to the substrate acetyl-CoA. The radiochemical procedure depends on enzymatic synthesis of (/sup 3/H)acetyl-CoA in the assay using (/sup 3/H)acetate, ATP, CoASH, and acetyl-CoA synthetase. NAT activity is measured by quantifying N-(/sup 3/H)acetylarylamine after separation from (/sup 3/H)acetate by extraction. Product inhibition by CoASH is prevented in this system by the use of acetyl-CoA synthetase.

• Research Organization: Univ. of Michigan Medical School, Ann Arbor
• OSTI ID: 5253617
• Journal Information: Anal. Biochem.; (United States), Vol. 2
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
TRANSFERASES
ENZYME ACTIVITY
ACETATES
AMINES
BIOASSAY
CATALYSIS
LIGASES
LIVER
MICE
RABBITS
RADIOCHEMISTRY
SPECTROPHOTOMETRY
TRACER TECHNIQUES
TRITIUM COMPOUNDS
ANIMALS
BODY
CARBOXYLIC ACID SALTS
CHEMISTRY
DIGESTIVE SYSTEM
ENZYMES
GLANDS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MAMMALS
ORGANIC COMPOUNDS
ORGANS
RODENTS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques