The mechanism of ethanol action at serotonergic neuronal systems in the brain was investigated by examining the effects of alcohols on the activity of tryptophan hydroxylase (TPH) in vitro using natural type of biopterin as cofactor. Alcohols inhibited the activity of TPH prepared from rat brain in a noncompetitive manner with respect to both the biopterin cofactor and the L-tryptophan substrate. The rank order of inhibitory potency of the tested alcohols was n-propanol > iso-propanol > ethanol > methanol. The kinetic study indicated that alcohols more potently affected the enzyme interaction with cofactor than substrate. Ethanol, at concentrations that can be reasonably attained in vivo (i.e., 25-100 mM) significantly decreased TPH activity in the presence of a physiological concentration of cofactor. However, the reduction was only approximately 5% of control activity, because Ki, values of ethanol for the enzyme were very high (800-1000 mM). From the present results, it was concluded that the direct inhibition of the synthetic enzyme itself by ethanol would contribute little to in vivo effects of ethanol on serotonergic neuronal systems.

1-Propanol↗

PubMedDate not supplied

Binding and uptake of glycoproteins to brain synaptosomes.

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Animals↗

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Alteration of the substrate specificity of Aspergillus oryzae beta-galactosidase by modification with polyethylene glycol.

beta-Galactosidase (beta-D-galactoside galactohydrolase, EC 3.2.1.23) purified from Aspergillus oryzae was modified with 2,4,6-trichloro-s-triazine derivatives of polyethylene glycol (activated BPEG) having molecular weights of 600, 1500, 2000, and 4000. Polyethylene glycol derivatives were attached to 6 of the 12 amino groups exposed on the surface of the enzyme. Upon modification, the enzymatic activity for a water-soluble substrate, o-nitrophenyl beta-D-galactopyranoside, was reduced with increasing molecular weight of the activated BPEG. On the contrary, the enzymatic activity for another substrate, 4-methylumbelliferyl beta-D-galactopyranoside, was increased upon modification. The Michaelis constants of native and modified enzymes for these two substrates were virtually the same. The effect of the modification was more marked in the enzymatic hydrolysis of the beta-galactosidic bond of amphipathic substrates. A fluorescent analog of naturally occurring galactocerebroside, 1-O-galactosyl-2-N-(1-dimethylaminonaphthalene-5-sulfonyl)-sphingosine, was hydrolyzed more rapidly by the modified enzyme than by the native one. The enzyme modified with activated BPEG of 1500 Da had the highest activity for this substrate. The beta-galactosidic bond of the terminal galactose of GM1-ganglioside (II3NeuAcGgOse4Cer, galactosyl-N-acetylgalactosaminyl-(N-acetylneuraminosyl)-galactosyl -glucosylceramide) was cleaved by the modified but not by the native enzyme.

Aspergillus↗