Abstract
A sensitive coupled assay for histidine decarboxylase has been developed. This method involved conversion of [3H]histidine into [3H]histamine by the enzyme sample, with methylation of this product in situ, catalysed by the enzyme histamine N-methyltransferase, to yield [3H]N-tele-methylhistamine. The radioactive product was separated from the substrate by (i) extraction into chloroform, (ii) ion-exchange chromatography and (iii) liquid cation-exchange extraction. The "no tissue" assay blank comprised 0.0007% of the substrate radioactivity. Sample material with a histidine decarboxylase activity of as little as 0.14 fmol/min/ml (measured at 1 microM histidine) gave double the blank value. More than 50 assays could be performed in one day. This assay was used to determine the in vivo changes in mouse brain histidine decarboxylase activity following irreversible inhibition with (+) alpha-fluoromethylhistidine (alpha-FMH). From the time course of recovery of enzyme activity the half-life of histidine decarboxylase in vivo was calculated to be 53 h.
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