Pig brain contains an enzyme that oxidizes the physiologically important aldehydes 5-hydro-xyindole-3-acetaldehyde, 4-hydroxy-3-methoxy-and 3,4-dihydroxyphenylacetaldehydes, d-3,4-dihydroxyphenylglycollaldehyde as well as d- and l -p -hydroxyphenylglycollaldehyde [A1287].
Rat liver enzyme is found in mitochondria (three isozymes, pI 5.4, 5.6 and 6.9) and cytosol (five isozymes, pI 5.8, 6.05, 6.15, 6.6 and 7.4). All of them oxidize p -nitrobenzaldehyde and 3,4-dihydroxyphenylacetaldehyde [B659]. Cytosolic enzyme oxidizes phenylacetaldehyde, and is induced by phenobarbital, DDT, polychlorobiphenyls and other xenobiotics [A2973]. Another study found three peaks of activity towards m -nitrobenzaldehyde. One isozyme required NADPH and a second required NADH. Both were identified as E.C. 188.8.131.52, the second being a 3a-hydroxysteroid dehydrogenase. The third required NADH, and was identified as alcohol dehydrogenase, E.C. 184.108.40.206 [A1688].
Cucumber enzyme oxidizes indole-3-acetaldehyde to indole-3-acetate. It is a metalloflavoprotein that does not require cofactors [A3378].
Achromobacter euridice phenylacetaldehyde dehydrogenase (E.C. 220.127.116.11), optimum pH 8.9, requires NAD+ (NADH does not catalyze the reverse reaction) and a monovalent cation (K+ is best). The reaction is irreversible and is highly specific; there is some action with indole-3-acetaldehyde, but other aldehydes show poor activity. It is unstable, but it is protected by substrates or 10 per cent acetone [K869].
Acinetobacter calcoaceticus benzaldehyde dehydrogenase II has a molecular weight of 51 654, with 484 amino acid residues, based on nucleotide sequencing [J621]. It is a tetramer, optimum pH 9.5 [E596]. The reaction is not reversible [E473].
Dehydrogenase I, which is involved in mandelate metabolism, is a tetramer, subunit molecular weight 56000, optimum pH 9.5 and pI 5.5. It requires NAD + ; NADP+ is less effective. It oxidizes a large range of benzaldehydes. There is an associated esterase activity with p -nitrophenyl acetate as substrate [F366].
A Flavobacterium constitutive benzaldehyde dehydrogenase requires NAD + , whereas a dehydrogenase induced by phenylglycine requires phenazinemethosulphate [E355].
Pleurotus eryngii aryl alcohol dehydrogenase oxidizes benzaldehydes and cinnamaldehydes, but mostly at a slow rate [G668].
Pseudomonas coniferaldehyde dehydrogenase, apparently a homodimer, molecular weight about 86000 and optimum pH 8.8, requires NAD + . Other substrates are trans-cinnamaldehyde, sinapaldehyde and benzaldehyde, but not vanillin [J707].
Pseudomonas putida produces two different aldehyde dehydrogenases depending on whether it is grown on p -cresol or 3,5-xylenol. The former enzyme is stable at 4°, but the latter enzyme is somewhat unstable at 4°; stability is improved in 10 per cent ethanol [A30]. Rhodopseudomonas acidophila contains two aldehyde dehydrogenases, one of which preferentially acts on aliphatic, and the other on aromatic aldehydes. The latter is a dimer, molecular weight about 70000, optimum pH 9.0 and pI 4.74. It oxidizes cinnamaldehyde and a series of benzaldehydes [G246].
Streptomyces aldehyde oxidase, molecular weight about 80 000, oxidizes vanillin [F225].
Benzaldehyde dehydrogenase I has been found in a Bacterium [A730].
Streptomyces viridosporus enzyme oxidizes vanillin and a range of benzaldehydes, but is inactive towards phthalaldehyde and aliphatics. It requires oxygen; peroxide is a second product [C284].
Citrus enzyme, molecular weight 200 000 and optimum pH 7.5, forms peroxide as well as indole-3-carboxylate; it is highly specific [H80].
Pea indole-3-acetaldehyde oxidase is composed of two isozymes that are not activated by pyridine nucleotides. The main has optimum pH 4.5, and the other optimum pH 7.0, with 3-formylindole as another substrate. It is not a dismutase [G446, G520]. Another report states that the optimum pH is 8.0; it requires oxygen [B861].
3.4 Reductions of acids
Aryl-aldehyde dehydrogenase (E.C. 18.104.22.168) a. Ar.COOH^ Ar.CHO
Clostridium formicoaceticum reductase, a dimer, monomeric molecular weight 67 000, contains iron and tungsten. It reduces a range of benzoates [G416].
Neurospora crassa enzyme has an optimum pH of 7. Benzoate initially yields benzoyl AMP, which then produces benzaldehyde. In the presence of hydroxylamine, benzoate yields benzoylhydroxylamate. Many benzoates and cinnamates are substrates, but a large proportion of these are at best very poor substrates.
The reaction scheme suggested is:
Aryl acid + Mg2+ + ATP i pyrophosphate + enzyme.acyl AMP
Enzyme.acyl AMP + NADPH i aldehyde + AMP + NADP+
Nocardia enzyme, molecular weight 140 000 or 163 000 (depending on the measurement method), reduces benzoate, and requires ATP and NADPH [H955]. N. asteroides enzyme, a monomer, molecular weight 152000, requires ATP, NADPH and Mn2 + ; benzoyl AMP is an intermediate in the reduction of benzoate. The enzyme acts on substituted benzoates, preferentially on meta-substituted acids, but usually not on those with ortho substituents. Some longer chain acids are also substrates [G207].
b. Ar.COOH^ Ar.CH2OH
Rat brain enzyme reduces dopac to 3,4-dihydroxyphenylethanol, optimum pH 7.5; it is unclear whether an alcohol dehydrogenase is part of the system. It is activated by Zn2 +, Mn2 +, Co2+ and Cu2 + , and EDTA is inhibitory. 3,4-Dihydroxymandelate, 4-hydroxy-3-methoxymandelate and 4-hydroxy-3-methoxyphenylacetate are not substrates [H798].
Cinnamoyl CoA reductase (E.C. 22.214.171.124)
Eucalyptus gunnii enzyme (cinnamoyl CoA: NADP+ oxidoreductase), molecular weight 38 000 and pI 7, acts on CoA conjugates of ferulic, sinapic and o -coumaric acids [H330].
Glycine max enzyme, molecular weight 38 000 and optimum pH 6.1, requires NADPH. Substrates include CoA conjugates of ferulic, sinapic, p -coumaric, 5-hydroxyferulic, caffeic and cinnamic acids. It is inhibited by thiol-binding reagents [A2576]. Spruce and Glycine enzymes are similar; in another study they were found to be dimeric, with the molecular weight of the native enzymes about 70 000. The reaction is reversible [B922].
Populus euramericana enzyme, a homodimer, monomeric molecular weight 40000, pI 7.5, is found in the stem. Substrates include CoA conjugates of ferulic, sinapic and p -coumaric acids, with NADPH as co-substrate [D54].
Flavobacterium enzyme, which requires phenazinemethosulphate as cofactor, acts on a number of d-amino acids, but not on the l-isomers. The product formed from d-phenylglycine is phenylglyoxylate [E355].
Mouse enzyme acts on phenylalanine and tryptophan [C219]. In one mouse strain some animals are devoid of activity; the genetics demonstrate the involvement of an inactive allele [C219a].
Pig kidney enzyme has a monomeric molecular weight of 38 000 or 39 600 using different methods [A151].
Rat kidney enzyme acts on d-dopa, and is inhibited by d-alanine and benzoate. The appearance of 3,4-dihydroxyphenylpyruvate predominates over formation of dopamine in kidney (presumably formed by reverse transamination of 3,4-dihydroxyphenylpyruvate followed by decarboxylation), but only at high substrate concentrations [A1393].
Studies on human, African green monkey, rat, pig and mouse kidney found activity with phenylalanine and tryptophan, but in chicken and frog the tryptophan activity was missing [C114].
Fish enzymes (electric eel, rainbow trout, carp, crucian carp and catfish) act on phenylalanine but not on tryptophan [C219].
These enzymes are an important component of snake venoms, and their concentration is sufficiently high in some to enable the raw venom to be used as a reagent for measuring specific amino acids.
Cerastes vipera oxidase is a dimer containing FAD, molecular weight 122000, monomeric molecular weights 61 000 and 64000 and optimum pH 7.5; phenylalanine is a substrate. It is activated by Cu2+ and Mn2 + , and inhibited by EDTA [G417].
Trimeresurus (Taiwan habu snake) venom enzyme is dimeric, molecular weight 140000 and pI 5.4, and contains two mol FMN/mol. Substrates are phenylalanine and tyrosine [E770]. Another study found pI 8.4 for habu snake enzyme. During purification the specificity changes, suggesting that there are several isozymes [A636].
Viper palaestinae enzyme is composed of three isozymes, molecular weight 130000 and optimum pH 8.8 with kynurenine as substrate [A1044].
Neurospora crassa enzyme, molecular weight 300 000, and optimum pH 9.5 probably contains four FAD/mol; phenylalanine is a substrate [A404].
L-Phenylalanine dehydrogenase (l-phenylalanine: NAD+ oxidoreductase, deaminating, E.C. 126.96.36.199)
l-Phenylalanine + NAD+ i phenylpyruvate + NH++ + NADH
Sporosarcina ureae and Bacillus sphaericus enzymes have been crystallized. They are both probably homooctamers, molecular weights 305 000 and 340000, pI 5.3 and 4.3, optimum pH 10.5 and 11.3 (oxidation), and 9.0 and 10.3 (reduction) respectively. S. ureae enzyme acts on phenylalanine (best) and on tyrosine, tryptophan, phenylalaninamide, phenylalanine methyl ester and l-phenylalaninol. B. sphaericus acts on phenylalanine and tyrosine, but poorly if at all on the other foregoing compounds [E445]. S. ureae enzyme aminates pyruvates [E423]; its molecular weight has also been reported to be 290 000 [D919].
Microbacterium enzyme, a homooctomer, monomeric molecular weight 41 000 and pI 5.8, requires NAD+ [J534].
Thermoactinomyces intermedius enzyme is a hexamer, monomeric molecular weight 41 000, that acts reversibly on l-phenylalanine [G222].
Nocardia enzyme is monomeric, molecular weight 42 000 and optimum pH 10 with phenylpyruvate, p -hydroxyphenylpyruvate and indole-3-pyruvate as substrates; other pyruvates are not substrates. The enzyme does not act on d-phenylalanine or other amino acids [F394].
Rhodococcus enzyme, optimum pH 10.1 for the forward reaction and 9.25 for the reverse, acts on phenylalanine (best), tyrosine and tryptophan and the corresponding pyruvates [E451]. It requires NAD+ or analogues; NADPH and analogues are inactive. The reaction sequence is binding of NAD + prior to phenylalanine; the products are released in the order ammonia, phenylpyruvate and NADH [K558].
Bacillus sphaericus enzyme acts on several phenylpyruvates with para substituents, and phenylpyruvate analogues with different side chain lengths. The products are the corresponding amino acids, which are formed quantitively [G168].
L-Tryptophan dehydrogenase and 2',3'-dioxygenase
Activity has only been found in one out of many Brevibacterium strains tested [D90].
L-Tryptophan dehydrogenase and 2',3'-dioxygenase (E.C. 188.8.131.52)
Pea (optimum pH 8.5), maize and tomato enzymes require NAD(P) for the reversible formation of indole-3-pyruvate [G483]. It is found additionally in Prosopis juliflora (mesquite) and wheat, but not in Brassica [D968].
Chromobacterium violaceum l-tryptophan 2',3'-dioxygenase forms 2',3'-dehydrotryptophan as the initial product, which can then isomerize to the imine, from which indole-3-pyruvate is formed by hydrolysis. The reaction, which is specific for the l-isomer, requires an unmodified indole nucleus and a carboxyl group. It also acts on some peptide hormones that contain tryptophan residues as well as other tryptophan analogues [H664]. The enzyme is polymeric, molecular weight 68 0000, with monomeric molecular weights 74000 and 14000, pi 4 and a broad optimum pH 3-8. It is a haem-containing mixed function oxidase, which forms peroxide from oxygen. N-substituted tryptophans (which cannot isomerize) form 2',3'-dehydro analogues of the substrates. Tryptamine, N-substituted tyrosine and phenylalanine are not substrates [H248]. Reaction with N-benzyloxycarbonyltryptophan involves a syn elimination [A2770].
In Pseudomonas N-acetyl-l-tryptophanamide forms N-acetyl-2',3'-dehydrotryptophanamide in two steps, the first of which is enzymatic, and forms 5-(3-indolyl)-2-methyl-2-oxazoline-4-carboxamide. This then forms 2',3'-dehydrotryptophanamide non-enzymatically with a time lag; at a lower pH, b-hydroxytryptophanamide is formed. The latter compound is further oxidized to b-oxotryptophanamide [A3617, C511]. Other products are indole-3-glycolaldehyde and probably indole-3-glyoxal [A3009].
These enzymes play an important part in the physiological control system for the catecholamine neurotransmitters in animals. Studies on these enzymes have led to the development of drugs that affect the amount of catecholamine available for neurotransmission, and, in consequence, valuable treatments for neurologically-induced illness have been developed as a result of the enormously extensive studies on this enzyme system. Of the many thousands of studies on monoamine oxidases it is possible to include in this review only a few that address basic enzyme properties.
a. Mitochondrial MAO.
MAO activity, which is associated with the inner mitochondrial membrane, is separated into two forms, distinguishable by inactivation by low concentrations of clorgyline at 10 ~8 M (MAOA) and deprenyl at 10~6 M (MAOB); these are 'suicide' inhibitors for MAO. In general, serotonin is oxidized by MAOA, and phenethylamine by MAOB [A1613, A1909, A2727, A2588]. Neither inhibitor is cleanly associated with MAOA or B; both inhibitions overlap in a concentration-dependent manner. Both clorgyline and pargyline (also an A inhibitor) and deprenyl initially inhibit reversibly, and this is followed by an irreversible phase. The conversion rate of clorgyline-inhibited MAOB (rat liver) into irreversibly inactivated enzyme is much slower than for MAOA. Deprenyl-inhibited MAOB is slowly converted into an irreversibly inactivated form, whereas there is no irreversible inactivation of MAOA by deprenyl [A1670, C46].
After solubilising human frontal lobe MAO with octylglucoside, MAOA and B were separated by chromatography on DEAE-Sepharose CL-6B [C682].
Studies using dopamine specifically labelled with deuterium in the a-position have demonstrated that the R - and not the S-deuterium is removed [D606]. a,a-D2-Phenethylamines (phenethylamine, tyramine and m-tyramine) are oxidized by rat liver microsomal enzyme at 25% of the rate for unlabelled compounds; the P,P-D2-analogues show a slight enhancement in activity [B660].
Kinetic studies suggest that the substrate amino group hydrogen bonds to an amino group in the enzyme. Introduction of a b-or p -hydroxyl group leads to a sharp drop in binding free energy, accompanied by a shift from B to A type substrates, possibly with a shift in orientation of the bound substrate [A1957].
Human MAOB acts on phenethylamine, dopamine and tyramine [F754]. By using clorgyline to inhibit MAOA and deprenyl to inhibit MAOB, studies on liver, kidney and cerebral enzyme have found that serotonin and 3-methoxytyramine are substrates for MAOA and phenethylamine for MAOB, whereas dopamine, tyramine and tryptamine are substrates for both MAOA and B [A2967, E55].
Phenethylamine is oxidized by human lung MAOA and B, whereas serotonin only by MAOA [A3957].
Dopamine is oxidized mainly by MAOB in dopamine-rich human brain areas, whereas MAOA contributes significantly to dopamine oxidation in most other tissues [A3688].
Monoamine oxidase activity is almost unaltered from normal in a range of areas in postmortem human parkinsonian brain [A381].
Human placental MAOA oxidizes phenethylamine and serotonin [A3957], as well as kynuramine, MPTP and a large range of MPTP analogues substituted in the phenyl ring [F395].
Activity of human platelet enzyme, which is almost exclusively MAOB [A1901] fluctuates during the oestrous cycle, with a maximum about the time of ovulation, and then drops by about 23 per cent to a minimum 5-11 days later [A367]. Activity is evenly distributed throughout human heart [A756]. A particulate enzyme in blood, thought to be mitochondrial, has an optimum pH between 6 and 8.8 depending on substrate, apparently without correlation with A and B status [A2010].
Rat liver enzyme can be distinguished into two distinct activities by the action of clorgyline and aryl nitriles [A367]. It is inhibited by the thiol-binding compounds nitroprusside and 5,5'-dithiobis(2-nitrobenzoate). Neither arsenite nor thiols are inhibitory [A238]. Hydroxylamine rapidly inhibits activity towards tyramine and benzylamine within one minute, whereas after one hour the activity towards tyramine, but not benzylamine is increased [A 1760]. It migrates electrophoretically as several bands, but after treatment with perchlorate, which releases lipid without change in activity, only one band is found [A 1039]. Using tyramine as substrate Km for MAOA and B are different [B661]. Both MAOA and B oxidize tryptamine, N-methyl-tryptamine, serotonin and 5-methoxytryptamine, whereas N-methylserotonin, bufotenine and N,N-dimethylserotonin are selective for MAOA, and N,N-dimethyltryptamine is selective for MAOB [B653].
Another study found that rat brain MAOA is heat stable, whereas B is labile. A and B are partially separated by sucrose density gradient centrifugation [A1227, A1309, Al841]; more MAOB is found in high-density mitochondria [A1909]. Both A and B are fairly evenly distributed throughout brain [A2386]. Brain enzyme is inhibited reversibly by b-propiolactone and b-nitropropionate [A1800], by apomorphine [A2069] and atropine (also heart enzyme) [A1898]. The specificity of A and B are similar to human enzyme (above) [A1324]. Brain mito-chondrial enzyme has an optimum pH of 7.5 for dopamine and tyramine, 8.2 /8.5 for serotonin and tryptamine, and 9.1 for kynuramine. Similar results were obtained with beef brain mitochon-drial enzyme [A1285]. Brain MAOA oxidizes dopamine better than MAOB [A2484]; m- and p -tyramine are substrates for both isozymes, whereas o-tyramine is substrate only for MAOB [A3667]. The ratio of B to A in synaptosomal mitochondria is about 1/2 that for extrasynaptosomal mitochondria from whole brain; the ratio is lower in striatal mitochondria than in cerebellar, and in cortical mitochondria it is lower still [A2487]. One study, however, suggested that both isozymes are probably extraneuronal [A3619]. Rat skeletal muscle contains both MAOA and B; A is more heat labile than B [A1798].
Rat brain enzyme activity increases sixfold to the adult level in the first 10 days following parturition [A2490].
Spinal cord transection in rat has no effect on cord MAOA or B activity [A562].
3-(N-Cyclopropyl)-5-phenylethylamino-1,2,4-oxadiazole is specific inhibitor for rat liver MAOA. Substitution with methyl or chloro in the p -position of the phenyl ring does not alter selectivity. Other substrates substituted on the m -position partially or completely lose their selectivity for MAOA [A794].
Rat A and B arterial isozymes are partially soluble. After treatment with 6-hydroxydopamine 70 per cent of MAOA activity was lost, whereas B was unaffected [A1045]. Rat lung shows both MAOA and B activities [A2161, A3863]. Another study found MAOB in pancreas as well as in lung, liver, kidney and brain [A2835].
Rat placental MAO activity increases threefold from day 15 /20 of gestation, and then declines by 50 per cent at term [A2469].
Rat heart enzyme is mainly MAOB at three weeks post partum, a mixture of A and B at eight weeks and only A in the adult [A3665].
Studies on partially lysed MAOA and B from rat hepatoma indicate that the peptide chains are different in these isozymes [A3683].
In rat, a series of oxo analogues of phenyl-ethanolamines are converted into mandelic acids, with phenylglyoxals as the initial products. The reaction is much faster than with the corresponding phenylethanolamines [J287]. Further studies (Goodwin, B.L., Ruthven, C.R.J., unpublished) with human placental MAO showed that MAOA oxidizes these b-oxophenethylamines.
Beef liver MAOB acts on benzylamine, MPTP and a large number of MPTP analogues substituted on the phenyl ring [F395]. These results (and the results for human MAOA) suggest that MAO is involved in MPTP-induced parkinsonism, and if parkinsonism is caused by the action of an environmental amine, the involvement of MAOB might explain the claimed beneficial effects of deprenyl in parkinsonism; MPTP does not appear to be the enviromental toxin [K948].
A study on beef brain cortical enzyme, using clorgyline, pargyline, harmaline and deprenyl as inhibitors suggested that the inhibition pattern is too complex to be simply due to MAOA and B. A two-stage inhibition is postulated, involving a rapid reversible step at two different binding sites, followed by a slow irreversible phase [A2181]. Treatment of this enzyme with cyanoacetyl hydroxyethylhydrazide induced the ability to oxidize histamine, cadaverine and analogues; this was prevented by pre-treatment with clorgyline, but not with deprenyl [A19l6].
Guinea pig brain activity increases 2.5-fold in the first 35 days following parturition, to 50 per cent of the adult level [A2490].
In hamster the ratio of MAOB to A activity varies from tissue to tissue. The ratio in brain is about 0.03, and in heart, lung, liver and spleen it is higher, but less than 1. In rat, the ratio is also consistently less than 1 in these tissues, but the ratio between tissues is different from that found in hamster. In rabbit the ratio for the above tissues is close to 1 [A2588].
Japanese monkey platelet enzyme is mostly B, with three components, molecular weights about 60000, pI 5.5, 6.5 and 7.0, whereas liver enzyme has pI 6.5. They can be solubilized with Triton X-100 [F758].
Mouse lung mitochondria, after fractionation by sucrose density gradient centrifugation, are separated into fractions with more MAOB than A in the higher density mitochondria [A1895]. Enzyme activity in the eye decreased after day one post-partum, increased to a peak at day five and then fell to the adult level at 14 days [A946]. In brain, it increased from day one to the adult level at two weeks except in cerebellum, where a slight increase continued until after week six [A1105]. Using tryptamine as substrate, brain and liver enzymes are inhibited by a range of b-carbolines, especially harmine and harmaline [A2178].
Pig brain enzyme shows a time-dependent inhibition by several N-substituted propargylamines, although this study found that not all compounds with this structure are inhibitory [A74]. A pig liver mitochondrial enzyme that acts on benzylamine (a benzylamine oxidase?) is a polymer with molecular weight about 1 200 000 containing about eight Cu and subunit molecular weight 146000 [A1033]. Pig dental pulp membrane-bound enzyme (presumably mitochondrial) oxidizes a range of amines, particularly phenethylamine and tryptamine [B206]. Pig heart enzyme oxidizes serotonin, tyramine and benzylamine, it is inhibited by clorgyline, and is considered to be heterogeneous [A2587]. The development pattern differs from organ to organ. Liver enzyme activity falls by 40 per cent at birth, and then increases threefold to the adult level at one month. Kidney and spleen activities rise steadily from 10 days pre-partum until 70 days post-partum. Heart activity reaches a maximum near birth, and then declines by 80 per cent over two months. Brain activity declines slightly near birth, and then increases slightly to the adult level; adrenal enzyme shows a similar pattern [A1569].
Rabbit lung and brain enzymes are inhibited reversibly by imipramine, DMI and DDMI [A94]. The ratio of MAOA to B in heart, kidney, lung, liver, spleen and brain is about 1 [A2588], and both A and B are found in platelets [A2326].
Terbutaline and orciprenaline are not substrates for rat and human liver MAO [A724].
The ratio of chick CNS MAOA and B depends on site, using phenethylamine and serotonin as markers. The highest ratio is found in spinal cord and lowest in cerebrum [A2386]. Brain enzyme activity is detectable at 14 days incubation, and increases steadily until 2 days post-hatching [A1413].
Squid brain enzyme can be differentiated into A and B [E147].
In man, the enzyme is found in all vascular tissues, localized in smooth muscle [B537]. Serum activity is reduced in burns and cancer patients [A2965]. It appears to be present in lung and placenta; however, benzylamine oxidation is considered to be catalyzed by MAOB in liver [A3957].
Human plasma and rat lung enzymes are not inhibited by deprenyl at 10~4 M (both MAOA and B are inactivated at this concentration) or by clorgyline, but semicarbazide, procarbazine and carbidopa are inhibitory. It is found (high activity) in aorta and lung, with slightly less in colon, ileum stomach, portal vein and duodenum, with low activity in some other tissues, including serum. Rat enzyme also oxidizes dopamine and phenethylamine, but these are not substrates in man [A2835, A3690].
Beef aorta enzyme is separable into two fractions; the major is particulate (probably not mitochondrial), and the minor cytosolic. N-Methylbenzylamine is a substrate only for the particulate enzyme. Carbonyl-binding reagents, especially pargyline are inhibitory [A574]. It is proposed that the mechanism for beef liver enzyme is ping-pong, with an imine intermediate being hydrolyzed to ammonia and aldehyde, with reduced flavoenzyme reacting with oxygen to form peroxide [A967].
Pig plasma enzyme is a dimer, monomeric molecular weight 95 000. Water of hydration bound to enzyme Cu2+ rapidly exchanges [A1025]. Heart enzyme has a molecular weight of 97 000, the same as plasma enzyme. It, also, is inhibited by carbonyl-binding reagents, and contains two Cu/mol [J833].
Pig dental pulp enzyme (soluble) oxidizes benzylamine, tryptamine and (less well) tyramine, but serotonin and phenethylamine are very poor substrates. This distinguishes it from MAOA and B [B206].
Rat brain contains a soluble enzyme that oxidizes kynuramine, but is not inhibited by deprenyl or clorgyline; this may be benzylamine oxidase [C186].
Rabbit enzyme that oxidizes mescaline appears to be similar to benzylamine oxidase [A1666].
c. Microorganism MAO.
Arthrobacter globiformis contains a soluble, Cu2 +-dependent enzyme (E.C. 184.108.40.206) that oxidizes phenethylamine. The peptide chain contains a 2,4,5-trihydroxyphenylalanyl residue as its quinone; this acts as cofactor in the reaction. It is formed spontaneously from a specific tyrosyl residue by the action of Cu2 + [H298]. The enzyme is a homodimer, molecular weight 141 000, with optimal activity and stability at pH 6.5. The products are presumably aldehydes, with peroxide as a second product. Other substrates include 3-phenylpropylamine, tyramine, dopamine, octopamine, tryptamine and 4-phenylbutylamine, but not benzylamine or aliphatic amines; carbonyl-binding reagents are inhibitors [F626, H912].
E. coli contains a phenethylamine oxidase with the above quinone cofactor, which is formed spontaneously by incubation of the enzyme at 30° and pH 6, especially at low enzyme concentration [H422].
Micrococcus luteus enzyme is a homodimer containing FAD, monomeric molecular weight 49 000 by standard methods; DNA studies indicate a molecular weight of 49 100, with 443 amino acid residues. Substrates include tyramine, adrenaline, dopamine and noradrenaline. It is inhibited by reagents specific for MAOA and B [K420].
MAO is membrane bound in some bacterial species, but is not present in others. It has been detected in Klebsiella, Escherichia, Salmonella, Pseudomonas, Brevibacterium and Micrococcus, and is induced by tyramine. Substrates include tyramine, dopamine, and (except in Micrococcus) octopamine and noradrenaline [B225].
d. Unspecified enzymes.
Helix pomatia enzyme is found in crop and nervous system, and a little in the heart [A657].
Hymenolepis diminuta enzyme is membrane-bound. It oxidizes (in decreasing order) dopamine, adrenaline, noradrenaline, tryptamine, tyramine and octopamine, but not serotonin or benzylamine. Inhibitors include cupferron, a,a-dipyridyl, iodoacetamide, pargyline, nialamide and iproniazid, but not azide, hydroxylamine or semicarbazide [A3815].
Tetrahymena pyriformis MAO oxidizes tryptamine, dopamine and serotonin. The activity increases in the pH range 6.5-7.8 [A3131].
Spermine: oxygen oxidoreductase (deaminating; E.C. 220.127.116.11)
Sheep plasma enzyme acts on benzylamine and nuclear-substituted benzylamines, tyramine, tryptamine and serotonin [A535].
Rat brain enzyme, which has also been called ephedrine-neotetrazolium chloride reductase (neotetrazolium chloride is the hydrogen-receptor co-substrate), is mainly mitochondrial, but some is cytosolic, with a dialysable heat-labile cofactor; NADP+ activates it. Substrates include ephedrine, adrenaline, tryptamine, serotonin, tyramine and noradrenaline, optimum pH 7.5. It is inhibited irreversibly by cyanide, but not by MAO inhibitors such as iproniazid or pargyline; iron chelators, such as o -phenanthroline and a,a-dipyridyl, are inhibitory. The nature of the reaction is not specified in Chemical Abstracts, but it appears to act on the amino groups, probably to form carbonyl compounds [A1053, A2489, A2490, A2802].
An Alcaligenes faecalis enzyme called aromatic amine dehydrogenase acts on tyramine and a range of phenethylamines to form aldehyde and ammonia, but not peroxide. The protein chain contains a crosslinked tryptophan tryptophanylquinone group, which is part of the redox system [H367, H421].
Pseudomonas aromatic amine dehydrogenase (aralkylamine dehydrogenase, E.C. 18.104.22.168) is an inducible tetramer composed of two pairs of monomers, molecular weights 46 000 and 8000 and optimum pH 7.5 /8.0, depending on substrate (tyramine, serotonin, tryptamine, phenethylamine, benzylamine and, best, dopamine); the products are ammonia and aldehydes. Traces of iron and copper are found, but these may be artifacts. The amino acid composition has been determined [C517, K709].
Octopamine hydro-lyase (E.C. 22.214.171.124)
Pseudomonas aeruginosa enzyme is soluble and forms p -hydroxyphenylacetaldehyde [K709].
Arthrobacter synephrinum enzyme, optimum pH 8.0 is cytosolic. It converts p-sympatol into p -hydroxyphenylacetaldehyde and methylamine, apparently anaerobically. It requires Mg2+ or Ca2 + , and is further activated by thiols. Tyramine, N-methyltyramine and hordenine are not substrates [A2915].
Nocardia also demonstrates this activity [E615].
p -Dimethylaminomethylbenzylamine is oxidized by human kidney enzyme. A study on inhibitors revealed that a series of dimethylsulphonium and trimethylammonium compounds with a hydrocarbon chain consisting of up to 12 methylene units, trimethylsulphonium, tetramethylammonium and ammonium compounds as well as phenelzine are competitive inhibitors, whereas pargyline is an uncompetitive inhibitor. A series of isothiuronium and guanidinium compounds with a chain consisting of up to 12 methylene units, 5-methylisothiuronium, guanidine, methylguanidine, hydralazine, iproniazid, nialamide and other compounds are noncompetitive inhibitors [A1686].
An enzyme in pig kidney that is claimed to be diamine oxidase acts on p -substituted benzyl-amines and many aliphatic amines [A3693]; it may be identical with benzylamine oxidase (R. Lewinsohn, personal communication).
3.6 Oxidative removal of substituents on amino groups
Aminopyrine N-demethylase activity is lower in red-winged blackbird than in rat; this difference parallels lower levels of P450 in bird microsomes [A358]. In male rat, activity increases 20-fold in the first 10 weeks post-partum, most of which occurs in the first four weeks. In female rat, activity is three times higher at birth than in the male, and it increases a further threefold in the following five weeks [A736]. The results of another study on rat liver differed considerably from this in detail [A71]. In rabbit, the activity increases sharply from a low level at about 20 days post-partum, and then remains level or declines slightly to the adult level [A1980].
Aminopyrine is demethylated by catalase and cumene hydroperoxide by a free-radical mechanism. It is suggested that the active site for this reaction differs from the one involving hydrogen peroxide [B378].
d-Benzphetamine is demethylated as the preferred substrate by all six P450 fractions separated electrophoretically in one study on human liver [C798].
p-Chloro-N-methylaniline demethylase is found in chick embryo liver and human foetal liver, adrenal, brain and lung [B8l5].
N,N-Dimethylaniline is demethylated by rat P450, horseradish peroxidase, beef liver catalase, human placental and Glycine max lipoxygenases, lactoperoxidase and haemoglobin, whale myoglobin and chloroperoxidase. Using substrate labelled with deuterium on one methyl group an isotope effect is observed with all these enzymes, the extent of which is specific for each enzyme. High isotope effects found with most of the haemoproteins tested are considered to indicate a rate-limiting step of hydrogen atom abstraction from the a-carbon (in this case defined as the methyl group), whereas deprotonization of the a-carbon is postulated for a low isotope effect, as observed with P450 and chloroperoxidase [C814]. Lipoxygenases also act on a range of other N-methylated anilines, with the formation of formaldehyde, but without any evidence for the formation of N-oxides as intermediates. Glycine enzyme has an optimum pH of 6.5 [K419].
N-Ethyl-N-methylaniline is dealkylated to both monodealkylated products. NADH enhances dealkylation, but not N-oxidation; dealkylation of the corresponding N-oxide is very feeble. These
Demethylation of N-methylated amides results, combined with the effects of selective inhibitors indicate that most of the dealkylation occurs directly, rather than via the N-oxide [A310].
Ethylmorphine N-demethylase is found in quokka, kangaroo, bettong, bandicoot, and possum (marsupials), as well as in rat [A2420]. Activity of rat and mouse liver enzymes is increased by EDTA and decreased by Fe2 + ; this probably correlates with inactivation by lipid peroxides [A89].
Imipramine demethylase activity found in rat hepatocyte microsomes is suppressed by SKF 525-A, a P450 inhibitor [K168]. Glycine lipoxygenase catalyzes demethylation, with peroxide as oxidant; apparently a free radical is an intermediate, and formaldehyde is released. A number of imipramine analogues are also demethylated [K359].
Human haemoglobin can dealkylate N-methylaniline and benzphetamine [D135].
N-Methylephedrine is demethylated by rat liver microsomes. The enzyme is inhibited by CO or SKF 525-A, but not by methimazole or by pre-incubation [F501].
Rat liver P450 N-demethylates 15 different compounds, utilizing either O2 and NADPH, or hydrogen peroxide. There is no correlation between the reaction rates corresponding to these mechanisms for the different compounds used; it is therefore considered that these are independent reactions [C669].
In rat, guinea pig and mouse N-demethylase activity is correlated with P450 activity [A627].
With some tertiary amines demethylation occurs in two stages, with the amine N-oxide as intermediate. This is described under the enzymes involved.
Rat liver microsomes demethylate N,N-dimethylbenzamide to form N-methylbenzamide and formaldehyde, with the corresponding N-hydroxymethyl compound as an intermediate. Several para-substituted
N,N-dimethylbenzamides are also substrates [G683].
Human bifunctional peptidylglycine a-amidating monooxygenase requires ascorbate, copper and oxygen, with hippurates as substrates. Salicyluric acid initially forms N-salicyl-a-hydroxyglycine, which then forms salicylamide and glyoxylate [K660].
Lipoxygenase (E.C. 126.96.36.199)
Glycine max enzyme, optimum pH 6.5, dealkylates aminopyrine by a free radical reaction, forming formaldehyde and peroxide. This is claimed to be a novel reaction [J526].
Pig liver enzyme, optimum pH 7.5, requires NADP; NADPH is ineffective. Tetrahydrofolate is formed, with the release of carbon dioxide from the formyl moiety [K877].
Pseudsomonas acidovorans enzyme, named pteridine 6-methylaminohydrolase, acts on pteroate or folate (it is unclear which compounds were tested) to cleave the C /N bond linking the pteridine and aminobenzoate moieties. Structural considerations indicate that this is an oxidative reaction, with formation of p -aminobenzoate [A971].
Dealkylation of quaternary bases
N-Methylnaltrexone is demethylated by rat, dog, man and mouse [F486].
Oxidation of amino to nitro groups
Man and monkey debutylate bupivacaine, but the reaction was not further studied [A255, J881, K14].
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