Adenylic deaminase and

The purine nucleotide cycle also is involved in muscle energy production. During intense stimulation, or when O2 supply is limited, the high-energy bond of ADP is used to synthesize ATP via the myokinase reaction (Figure 21-12). The resulting AMP can dephosphorylate to adenosine, which diffuses out of the cell. Conversion of AMP to IMP via adenylate deaminase and then to adenylosuccinate helps sustain the myokinase reaction, especially in FG fibers, by reducing accumulation of AMP. It may also reduce the loss of adenosine from the cell, since nucleosides permeate cell membranes while nucleotides do not. 

Fishbein, W.N., J.L. Griffin, K. Nagarajan, J.W. Winkert, and V.W. Armbrustmacher. 1979. Immunologic uniqueness of muscle adenylate deaminase and genetic transmission of the deficiency state. Clin. Res. 27 274A. 

Candida utilis is grown to high biomass concentrations and the extracted RNA is subsequently hydrolysed into the four 5 nucleotides adenosine 5 -monophosphate (AMP), GMP, cytidine and uridine 5 -monophosphate by crude nuclease PI from Penicillium the desired nucleotides are isolated by ion-exchange chromatography and AMP is converted to IMP by adenyl deaminase from Aspergillus [22, 36]. 

Purine nucleotides are degraded by a pathway in which they lose their phosphate through the action of 5 -nucleotidase (Fig. 22-45). Adenylate yields adenosine, which is deaminated to inosine by adenosine deaminase, and inosine is hydrolyzed to hypoxanthine (its purine base) and D-ribose. Hypoxanthine is oxidized successively to xanthine and then uric acid by xanthine oxidase, a flavoenzyme with an atom of molybdenum and four iron-sulfur centers in its prosthetic group. Molecular oxygen is the electron acceptor in this complex reaction. 

It has long been known that the adenylic deaminase of blood is increased in shock (Tl). Seligson s experiments with animals in hemorrhagic shock demonstrated that the peripheral blood ammonia was markedly elevated (N4). Further work with this problem showed that the blood ammonia rises during hemorrhagic shock to tremendous levels and that these levels are compatible with the cerebral symptoms noted in shock (H2). Retransfusion of the bled animal does not cause this ammonia level to return completely to normal, and in fact it remains elevated to toxic levels until the death of the animal. A source of this ammonia has been shown to be the intestinal tract, for the highest rise of ammonia is found in the portal system. 

S -Phosphodiesterase Several reports have appeared describing the semi-industrial scale application of immobilized 5 -phosphodies-terase for production of S -mononucleotides (IMP, AMP, GMP, UMP, CMP) to be used as flavor enhancers (30,41,42). In Japan, the enzyme is immobilized on a porous ceramic support and used (in combination with a similarly immobilized 5f-adenylate deaminase to convert AMP to IMP) to produce the S -mononucleotides from RNA (30,41). The deamination is desirable since IMP and GMP act synergistically with monosodium glutamate as a flavor enhancer. 

Figure 9.95 HPLC elution profiles of an adenylate deaminase incubation mixture. The reaction mixture contained 15 /xmol of imidazole-HCl (pH 6.8), 250 nmol of FoMP, 250 nmol of ATP, and 5 /u,mol of KCl in a final volume of 250 fiL. The reaction was initiated by the addition of activity obtained from the S-100 fraction and incubated at 37°C. At 10-minute intervals, 25 fiL samples were injected onto the HPLC column. There is a decrease in the FoMP peak (retention time, 1.7 min) and a significant rise in the peak corresponding to FoIMP (retention time, 3.1 min) up to 30 minutes. Inset Graphical representation of the first 30 minutes of the reaction. (From Jahngen and Rossomando, 1984.)...

Figure 10.10 AMP was formed from adenosine and ATP in a reaction catalyzed by adenosine kinase. After a 10-minute incubation, adenylate deaminase was added to the reaction mixture, and samples were taken and analyzed by HPLC Samples were analyzed every S minutes.

Due to the importance of modified nucleosides that are also in the base, the development of new biocatalytic processes appUed to the synthesis of derivatives modified in the base is of great interest Adenosine deaminase (ADA) and adenylate deaminase (AMPDA) are biocatalysts that catalyze the hydrolytic deaminahon of purine nucleosides and nucleotides. Some applications of these deaminases for the preparation and transformation of compounds structurally related to nucleosides with potential antitumor and antiviral activities have been described in the last few years [8]. 

Amino acids ate not the only source of ammonium ions produced in the body. Much of the ammonia produced, especially in the brain, arises from the hydrolysis of purines. Adenylate deaminase catalyzes the hydrolysis of AMP, yielding IMP and ammonium ions Cooper and Plum, 1987). IMP is inosine monophosphate (inosinic acid), GMP may also be hydrolyzed in this manner, yielding xanthosine and ammonium ions. Further details on purine metabolism occur at the end of this chapter and under Folate in Chapter 9. 

Lack of muscle adenylate deaminase has been found in some muscle disorders in which other abnormalities could not be identified. Failure to produce ammonia during intense effort may be diagnostic. In the heart, AMP accumulation is typically due to ischemia, and the adenosine released as a result is a potent coronary vasodilator. Accordingly, myocardium has less adenylate deaminase than skeletal muscle. However, large losses of adenosine from myocardium are dangerous because they lead to a decrease in ATP concentration that does not respond to dilation, thrombolytic, or oxygen therapy. Lack of adenosine deaminase in lymphoid tissue causes a severe immunodeficiency (Chapter 27). 

Myosin sufficiently purified in the usual way does not dephosphorylate any of the numerous phosphate compounds of living muscle other than ATP, and perhaps ITP, nor can it transphosphorylate. Menne (1943) finds that myosin, unlike the other main fractions of muscle, can convert arginine, histidine, glycocyamine, and choline into creatine. The myosin used, however, was only reprecipitated once and subsequently washed, and it is possible that the enzyme activity might be lost on further precipitation. After fractionation and precipitating three times, myosin possesses an appreciable adenylic deaminase activity (Hermann and Josepovits, 1949 Summerson and Meister, 1944). 

Much of the inosinate that is not synthesized de novo is formed via adenosine, as outlined above. Some inosinate is produced via adenylate deaminase. The significance of this enzyme in priming the Krebs cycle has been emphasized by Setlow and Lowenstein (S6). The IMP formed can be reconverted to either AMP or GMP. The deaminase is under control of ATP and GTP (B20). Thus this route may be of importance in maintaining the A G ratio in the cellular nucleotide pool. 

B20. Burger, R, and Lowenstein, J. M., Adenylate deaminase. III. Regulation of deamination pathways in extract of rat heart and lung. J. Biol. Chem. 242, 5281 (1967). 

Synthesis of modified purine nucleosides and related compounds mediated by adenosine deaminase (ADA) and adenylate deaminase (AMPDA) 05S509. 

An important discovery, that of free adenylic acid in muscle, was made by Embden in 1927. Muscle adenylate was recognized as the 5 -mono-phosphoric ester of adenosine because enzymatic deamination yielded the known inosinic acid. It was shown at that time that the deaminase preparations from muscle did not deaminate the adenylic acid isolated from alkaline hydrolysates of yeast nucleic acid as well, differences were apparent in the chemical properties of the adenylic acids from these two sources. Yeast adenylic acid and the other nucleotides from alkaline hydrolysates of RNA were ultimately shown to be mixtures of the 2 - and 3 -phospho esters. In 1929 the isolation of adenosine triphosphate from muscle was reported by Lohmann and independently by Fiske and Subbarow. The discovery of adenosine diphosphate followed in 1935. 

Increased enzyme activity may be due to increased concentrations of nucleotide substrates, or by other mechanisms. For example, GTP is required as a substrate for adenylosuccinate synthetase, and ATP is required for guanylate synthetase in addition, ATP activates adenylate deaminase (33). 

Adenylate deaminase is inhibited by GDP and GTP (37-40) and these inhibitors have mutually antagonistic effects toward the activator ATP (40). The situation is complicated by enzyme activation by Na+ or K+, and in some (but not all) preparations these cations prevent the stimulatory effect of ATP. These phenomena are summarized in Table 9-1. 

Guanylate reductase, which deaminates this nucleotide, catalyzes a reductive, rather than hydrolytic, deamination and has been discussed in Chapter 9. Like adenylate deaminase, it has a catabolic role and also functions in purine nucleotide interconversion. A guanosine deaminase has recently been identified in a pseudomonad (13), but it is not known to occur in animal cells. 

In rat heart the control rate of adenylate deaminase activity was lower, and that of adenylate dephosphorylation higher than in lung. Most of the ammonia formed from adenylate was therefore due to adenosine deaminase activity. ATP stimulated adenylate deaminase to almost the same relative degree in heart as in lung, but due to a marked inhibition of dephosphorylation the total amoimt of ammonia formed was less in heart. These data also raise questions concerning the identity and substrate specificities of the enzyme(s) that dephosphorylate adenylate and inosinate. 

A 5 -adenylic deaminase found in muscle acts on 5 -AMP and 5 -dAMP. The affinity of the enzyme is much greater for AMP than for dAMP. The pH for optimum activity is 5.9. 

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