Measurement of folylpolyglutamate synthetase in mammalian tissues.
Previous methods for the measurement of folylpolyglutamate synthetase have been modified and combined to facilitate assay of this enzyme at the levels found in mammalian tissues. Batch adsorption of product onto charcoal allowed the rapid analysis of multiple samples of partially purified enzyme, e.g., column fractions. This technique, however, was unsuitable for the assay of folylpolyglutamate synthetase in crude cytosols due to the presence of interfering enzyme activities. On the other hand, the sequential use of charcoal adsorption and batch elution from DEAE-cellulose permitted isolation of the folate product from assay mixtures containing crude enzyme fractions. Under these conditions, interference from other enzyme activities and background values were low enough for the quantitation of 10 pmol of oligoglutamyl folate product. Folylpolyglutamate synthetase was measured in a series of mouse tissues and tumors. Enzyme activity was quite low in all cases. Mouse liver and kidney and some of the tumors studied had the highest levels (50-100 pmol product/h/mg protein); other tumors and spleen had lower levels. Enzyme activity was at the limit of detection in intestine and lung and was below detection in brain, heart, and skeletal muscle.
D000327:Adsorption; D000818:Animals; D002606:Charcoal; D002848:Chromatography, DEAE-Cellulose; D002851:Chromatography, High Pressure Liquid; D003600:Cytosol / Q000201:enzymology; D005260:Female; D007422:Intestines / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D051379:Mice; D008811:Mice, Inbred DBA; D009374:Neoplasms, Experimental / Q000201:enzymology; D010453:Peptide Synthases / Q000302:isolation & purification*; D051381:Rats; D011919:Rats, Inbred Strains
D002606:Charcoal; D010453:Peptide Synthases; C023103:folylpolyglutamate synthetase