The rapid and specific determination of picomole quantities of gamma-aminolevulinate has been accomplished by its enzymatic conversion to uroporphyrinogen I and subsequent fluorometric detection of the oxidized uroporphyrin I. The coupled-enzyme assay was linear with time and protein concentration and required less than 3 h for 25 individual determinations. Under the standard assay conditions, 5-100 pmol of uroporphyrin I was reliably quantitated; these values corresponded to a range of gamma-aminolevulinate synthase activities from 0.2 to 15 nmol/h/ml enzyme. The sensitivity of this method was comparable to the more time-consuming radiochemical determinations of gamma-aminolevulinate synthase activity. The gamma-aminolevulinate synthase activity of liver homogenates from uninduced rats was 8.6 U/g liver (37 degrees C).
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A novel mutation of the erythroid-specific gamma-Aminolevulinate synthase gene in a patient with non-inherited pyridoxine-responsive sideroblastic anemia.
Am J Hematol
October 1999
Department of Clinical Medicine, Tohoku University School of Medicine, Sandai, Japan.
A novel missense mutation, G663A, in exon 5 of the erythroid-specific delta-aminolevulinate synthase gene (ALAS2) was identified in a Japanese male with pyridoxine-responsive sideroblastic anemia. Activity of the mutant delta-aminolevulinate synthase protein expressed in vitro was 15.1% compared with the normal control, but was increased up to 34.
View Article and Find Full Text PDFThe rapid and specific determination of picomole quantities of gamma-aminolevulinate has been accomplished by its enzymatic conversion to uroporphyrinogen I and subsequent fluorometric detection of the oxidized uroporphyrin I. The coupled-enzyme assay was linear with time and protein concentration and required less than 3 h for 25 individual determinations. Under the standard assay conditions, 5-100 pmol of uroporphyrin I was reliably quantitated; these values corresponded to a range of gamma-aminolevulinate synthase activities from 0.
View Article and Find Full Text PDFWe describe here a standard protocol for the radiochemical assay of gamma-aminolevulinate synthase from 17-day chick embryo liver mitochondria and starved male rat liver mitochondria. The assay is sensitive to at least 15 pmol of the gamma-aminolevulinate produced and can be adapted to suit most tissue types. A number of possible modifications to the procedure are discussed.
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