Theodore A. Alston

The bioorganic chemistry of the nitroalkyl group

Abstract Nitroalkyl groups are conspicuously rare among pharmaceutical agents, and the bioactivity of substituted nitroalkanes has been described in few instances. This article examines the natural occurrence of such compounds and the reactions of nitroalkyl compounds with enzymes. The discussion is not limited to nitro hydrocarbons; the term “nitroalkyl” is intended to distinguish the compounds from nitroaromatic compounds, which are better known in pharmacology and involve different chemical considerations. Further study of the bioorganic chemistry of the nitroalkyl group may contribute to the understanding of biosynthetic strategies and enzymatic catalysis and may permit the rational design of useful bioactive molecules.

Inactivation of alcohol dehydrogenase by 3-butyn-1-ol☆

Abstract Horse liver and yeast alcohol dehydrogenases are rapidly inactivated during their catalysis of the oxidation of 3-butyn-1-ol. In the case of the horse liver enzyme, the inactivation is secondary to covalent modification of the apoenzyme by an electrophilic product that accumulates in the reaction solution and that can also react with water, glutathione, and other enzymes. The modified protein exhibits enhanced ultraviolet absorbance, which is not bleached upon dialysis of the denatured enzyme at pH 7.4 for 24 h. The inactivation by 3-butyn-1-ol is more rapid than that which is afforded by the related alcohols 2-propyn-1-ol and 2-propen-1-ol under identical conditions and no inactivation is seen upon incubation with 3-hydroxypropanoic nitrile plus nicotinamide-adenine dinucleotide.

Amino Acid Neurotransmitters in the CNS: Properties of Diaminobutyric Acid Transport

Abstract: Uptake of L‐2,4‐diaminobutyric acid (DABA), a positively charged analogue of γ‐aminobutyric acid (GABA), by a synaptosomal fraction isolated from rat brain occurred with a Km of 54 ± 12 μM and a Vmax of 1.3 ± 0.2 nmol/min/mg protein. The transport of DABA was inhibited competitively by GABA whereas that of GABA was affected in the same manner by addition of DABA. The maximal accumulation of DABA ([DABA]i/[DABA]e) was observed to increase as the second power of the transmembrane electrical potential ([K+]i/[K+]e) and the first power of the sodium ion concentration gradient. These findings indicate that DABA is transported on the GABA carrier with a net charge of + 2, where one charge is provided by the cotransported Na+ and the second is contributed by the amino acid itself. Since uptake of GABA, an electroneutral molecule, is accompanied by transfer of two sodium ions, the results obtained with DABA suggest that one of the sodium binding sites on the GABA transporter is in proximity to the amino acid binding site.

Inactivation of alanine aminotransferase by the neurotoxin β-cyano-L-alanine

Abstract β-Cyano-L-alanine inactivates pig heart alanine aminotransferase. The nitrile and enzyme form a freely dissociable Michaelis complex which rearranges to a form of inactive enzyme. The inactivated enzyme slowly recovers activity at 25° in 100 mM phosphate buffer, pH 7.4. The observations are consistent with a mechanism of inactivation similar to that thought to apply to the suicide inactivator propargylglycine except that the putative covalent modification of the apoenzyme is relatively labile in the case of the nitrile.

Inhibition of succinate dehydrogenase by nitroacetate and by the toxic antibiotic nitraminoacetate

Summary Nitraminoacetate and the nitronate of nitroacetate are effective inhibitors of beef heart succinate dehydrogenase, respectively binding about 17 and 6.6 times more tightly than succinate at pH 8.0 and 25° as judged by the K m /K i ratios. Unlike the case of the suicide inactivator 3-nitropropionate, these compounds are freely reversible inhibitors. The toxicity of nitraminoacetate, an antibiotic elaborated by Streptomyces noursei , may be secondary to the inhibition of the Krebs cycle at the succinate dehydrogenase reaction, and the nitramino group may prove useful as a carboxylate analog for other enzymes.

Inactivation of pyridoxal 5'-phosphate-dependent enzymes by 5-nitro-L-norvaline, an analog of L-glutamate

Substrate analogs bearing ionizable nitro groups in place of carboxylate groups have proven to be interesting inhibitors of enzymes [l-5]. For instance, 2nitropropane (which is isoelectronic with the alanine zwitterlon) and the toxic antibiotic 3-~tropropionate (which is isoelectronic with succinate) irreversibly inactivate the flavin-dependent enzymes D-amino acid oxidase and succinate dehydrogenase, respectively, in m~chanist~~~ly distinct ‘suicide’ reactions [3]_ The nitronates obtained upon deprotonation of aliphatic nitro compounds generally bind well to enzymes which act on corresponding carboxylic compounds. In fact, in special cases in which the nitronates function as ‘transition-state analogs’, they bind by orders of magnitude more tightly than the carboxylic substrates [4,5]. Here, we report that several pyridoxal 5’-phosphate-dependent enzymes are subject to inactivation by a nitro analog of their substrate ~utamate. Although 5-nitro-L-norvaline (fig.2) sterically resembles L-2-aminoadipic acid somewhat more closely than it resembles L-glutamic acid, the lower homolog of S-~tro-L-no~ali~e is an unsown and probably unstable [6] compound. However, the nitronate form of 5nitro-L-norvaline exhibits nearly the same intramolecular charge separation as occurs in the L-glutamate anionic species; and 5-nitro-L-norv~ine is accepted into the active sites of alanine aminotransferase (EC 2.6.1.2), aspartate aminotransferase (EC 2.61 .l), and 4-aminobutyrate aminotransferase (EC 2.6.1.19), all of which act on L-glutamate.

Oxidation of N-nitroethylenediamine, a GABA analog from Agaricussilvaticus, by GABA aminotransferase

Abstract N-Nitroethylenediamine is a mushroom product which closely resembles the neurotransmitter 4-aminobutyrate, GABA. The nitramine is sequentially accepted as a substrate by the GABA-catabolizing enzymes GABA aminotransferase (EC 2.6.1.19) and succinic semialdehyde dehydrogenase (EC 1.2.1.16). In view of the steric and ionic similarity of the nitramino group to the carboxymethyl group, nitramines may prove generally useful for enzymological and pharmacological purposes as analogs of carboxylic acids.

Neurotoxicity of metronidazole.

Excerpt To the editor: In their review of antibiotic neurotoxicity, Snavely and Hodges (1) mention that the neurotoxicity of isoniazid stems largely from antagonism of vitamin B6. However, they poi...

Enzymatic release of nitric oxide from L-alanosine, an antineoplastic antibiotic

Abstract L-Alanosine is an antineoplastic drug which is the 3-isonitramino analog of L -aspartic acid. The drug is known to be metabolized to the corresponding 2-oxo acid. Unlike the parent amino acid, the 2-oxo acid is unstable under mild conditions. When the 2-oxo acid is generated in vitro by the aerobic action of L -amino acid oxidase on L -alanosine, the reaction mixture contains products capable of diazotizing sulfanilamide and of reducing ferricytochrome c to ferrocytochrome c . It is thus likely that, as expected from model reactions, the unstable 2-oxo acid derived from L -alanosine decomposes into nitric oxide and other reactive free-radical species. Enzymatically promoted production of highly cytotoxic nitric oxide may pertain to the biological activity of the antibiotic. The reaction should prove extrapolable to the design of other enzyme-activated cytotoxic agents.

Inactivation of GABA Aminotransferase by 3-Nitro-1-Propanamine

3-Nitro-1-propanamine is a close structural analog of the neuro-transmitter GABA. The nitro compound is a good substrate for the GABA aminotransferase from porcine brain. However, it inactivates the GABA aminotransferase from GABA-grown Pseudomonas fluorescens in a slowly reversible reaction. Both enzymes are inactivated by the homolog 4-nitro-1-butanamine.