The concentrations of S-adenosyl-L-methionine (SAM), S-adenosyl-L-homocysteine (SAH), and adenosine (Ado) were determined in whole brain and rat brain regions by HPLC. The whole brain contains, respectively, 22 nmol, 1 nmol, and 65 nmol of SAM, SAH, and Ado per g of wet tissue. Their distribution indicated that SAM and SAH levels are highest in brainstem, whereas the Ado level is highest in cortex. With aging the SAM concentrations decrease in whole brain, brainstem, and hypothalamus (-25%) and SAH levels increase by 90% in striatum and by 160% in cerebellum, while Ado levels are increased in all regions by 100--180%.

The heterogeneity of histidine decarboxylase from rat gastric mucosa was studied. The partially purified enzyme was fractionated by preparative isoelectric focusing on a flat-gel bed by using narrow pH-range carrier ampholytes and a short focusing time. The activity was resolved, with about 95% recovery, into three forms, designated I, II and III, with pI values of 5.90, 5.60 and 5.35 respectively. These three forms exhibited similar molecular weights, indicating that the forms were not the result of different degrees of polymerization. By preparative refocusing each form refocused as a single peak of enzyme activity with reproducible pI, but a high loss of activity occurred with repeated focusing. Forms I, II and III were purified by the combined use of preparative isoelectric focusing and gel chromatography and other fractionation methods. The active forms could be distinguished by electrophoresis and isoelectric focusing on polyacrylamide gels and displayed protein heterogeneity. These forms were found in the crude extract and in the partially purified preparations in the presence or absence of proteinase inhibitors. Form II had the highest specific activity, but all three forms had the same optimum pH and Km value for histidine.

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The properties of specific histidine decarboxylase from highly purified rat gastric mucosa preparations were studied. The kinetic parameters were pH dependent: the apparent K, value varied inversely with pH; the maximum reaction velocity was reached at pH 6.6; the optimum pH was related to substrate concentration. The enzyme was unstable below pH 5.5. The effect of temperature was investigated and the enzyme activity was optimum near 56°C. The thermal inactivation of the enzyme showed the presence of several active forms displaying distinct thermostabilities. The effect of coenzyme and substrate on heat stability was established. A small amount of pyridoxal phosphate was required for maximum enzyme activity, and the K, was low. The cofactor appeared to be tightly bound to the apoenzyme; nevertheless there was a fraction more easily resolved by dialysis. With high pyridoxal phosphate concentrations non-competitive inhibition occurred. Histamine inhibited the enzyme at high concentrations, the inhibition being competitive with respect to the substrate. No metal ion was required for enzyme activity; the enzyme was inhibited by sulfhydryl reagents and heavy metal ions, and also by high concentrations of reducing agents. The tryptophan residue of the holoenzyme seemed to be essential for the catalytic process.Histamine, which has an important role in a variety of physiological and pathological responses [I], is synthesised through the decarboxylation of histidine. In mammalian tissues the synthesis can be catalyzed by an enzyme which has a high specificity for histidine and is different from the nonspecific aromatic-L-amino acid decarboxylase. This specific histidine decarboxylase is found in many tissues but its activity is generally low; however, fetal rat tissue, hamster placenta and mastocytomas have high activities [2 -51. A high histamine-forming capacity has also been found in the gastric mucosa of several mammalian species including man [6-111. Studies concerning the histidine decarboxylases isolated from different sources allowed us to establish that they were similar in many respects [12]. However, the gastric enzyme studies have been principally performed on crude or partially purified preparations. The fact that rat gastric mucosa, which is rich in specific histidine decarboxylase, also contains a high activity of the non-specific enzyme [6] results in a difficult and false characterization of the specific enzyme.The properties of gastric histidine decarboxylase can be studied with accuracy only after the enzyme has been purified. Attempts to purify the specific gastric enzyme have been limited by the enzyme lability upon fractionation [9,13]. However, in a previous study [I41 we reported that a high degree of purification of the specific histidine decarboxylase from rat gastric mucosa had been obtained; only the specific enzyme was obtained by that procedure, the non-specific enzyme was absent. Its physicochemical properties were investigated, the enzyme consists of three active forms, the stability and ...