Enzymes adenosine deaminase reaction

Pteridine is changed by, the enzyme adenosine deaminase to the levorotatory form of the hydrate, 3,4-dihydro-4-hydroxypteridine. When this reaction was approached from the other direction, the same enzyme dehydrated this enantiomer most rapidly, leaving a net positive optical rotation at equilibrium.65 Specimens of the enzyme from both mammalian and fungal sources, although of very different molecular weight, were found to catalyze the stereospecific hydration of pteridine also pteridine inhibited the deamination of adenosine by both enzyme specimens. Assuming that the first step in the deamination of adenosine is the formation of a tetrahedral hydrated intermediate, it was argued that both the hydration of pteridine and the deamination of adenosine were parallel phenomena.65... 

Adenosine is degraded in a two-step reaction. First, the enzyme adenosine deaminase acts on AMP or adenosine nucleoside to yield IMP or inosine. 

An assay for adenosine involves the primary enzyme adenosine deaminase, and a chemical indicator reaction that consumes ammonia by reaction with the ninhy-drin reagent (Eq. 3.4) ... 

Persons with severe combined immune deficiency (SCID) are totally unable to mount an immune response to antigens. Both the B and T lymphocytes are affected. The disease arises from an inherited lack of a degradative enzyme, adenosine deaminase (ADA). The reaction shown here illustrates the pathways affected. Lack of ADA allows deoxyadenosine triphophosphate (dATP) to accumulate from the degradation of DNA. High dATP levels inhibit production of the other dNTPs needed for DNA replication because of their allosteric effects on the enzyme ribonucleotide reductase. 

The activity of this enzyme appears to be in the range of 0.027 umolar units/ml of erythrocytes. Recently, Meyskens and Williams (1971) have reported that the for adenosine for this enzyme is 1.9 X 10 6 M. If adenosine were to enter the erythrocyte in relatively high concentrations (as during tissue breakdown), the degradative adenosine deaminase reaction would be expected to predominate. On the other hand, if adenosine entered the erythrocyte in relatively low concentrations, e.g., 1 x 10 M, as the result... 

The structure of the urease active center is similar to that of adenosine deaminase, an enzyme containing one zinc(II) per active site (though see 48). This enzyme catalyzes the deamination of adenosine to inosine and NH3 (see Scheme 9), a reaction mechanistically related... 

Compound 25 (Fig. 18.9), a prodrug of 9-P-D-arabinofuranosyl guanine (26), was developed for the potential treatment of leukemia. Compound 24 is poorly soluble in water and its synthesis by conventional techniques is difficult. An enzymatic demethoxylation process was developed using adenosine deaminase (Mahmoudian et al., 1999, 2001). Compound 25 was enzymatically prepared from 6-methoxyguanine (27) and ara-uracil (28) using uridine phosphorylase and purine nucleotide phosphorylase. Each protein was cloned and overexpressed in independent Escherichia coli strains. Fermentation conditions were optimized for production of both enzymes and a co-immobilized enzyme preparation was used in the biotransformation process at 200 g/L substrate input. Enzyme was recovered at the end of the reaction by filtration and reused in several cycles. A more water soluble 5 -acetate ester of compound 26 was subsequently prepared by an enzymatic acylation process using immobilized Candida antarctica lipase in 1,4-dioxane (100 g/L substrate) with vinyl acetate as the acyl donor (Krenitsky et al., 1992). 

Figure 4.4 The HPLC analysis of a reaction mixture containing AMP and alkaline phosphatase. Separations were carried out on a reversed-phase column with a mobile phase of potassium phosphate (pH 5.5) and 10% methanol. The column was eluted isocratically, and the detection was at 254 nm. Two sets of tracings were obtained, according to the following schedules. For the original reaction mixture (A) immediately after the addition of enzyme, (B) after 10 minutes, and (C) after 15 minutes. For the reaction mixture to which had been added EHNA (5 /xAf), an inhibitor of adenosine deaminase, the suspected contaminant (D ) after 2 minutes, ( ) after 10 minutes, and (F) after 40 minutes. (From Rossomando et al., 1981.)...

Figure 10.11 AMP can be formed by adenosine kinase (1) in a reaction that uses ATP as the phosphate donor and forms ADP as the second reaction product. Alternatively, AMP can be deaminated to IMP by the enzyme AMP deaminase (2) and converted to inosine (INO) by a 5 -nucleotidase activity (3). Finally, AMP can be phosphorylated to ADP by the enzyme AMP kinase (4).

The salvage pathway does not involve the formation of new heterocyclic bases but permits variation according to demand of the state of the base (B), i.e. whether at the nucleoside (N), or nucleoside mono- (NMP), di- (NDP) or tri- (NTP) phosphate level. The major enzymes and routes available (Scheme 158) all operate with either ribose or 2-deoxyribose derivatives except for the phosphoribosyl transferases. Several enzymes involved in the biosynthesis of purine nucleotides or in interconversion reactions, e.g. adenosine deaminase, have been assayed using a method which is based on the formation of hydrogen peroxide with xanthine oxidase as a coupling enzyme (81CPB426). 

Adenosine deaminase (ADA), which catalyzes the conversion of adenosine to inosine (Equation 17.42), is an extremely proficient enzyme, providing a rate enhancement of more than 12 orders of magnitude (123). The enzyme-catalyzed reaction is thought to pass through an unstable hydrated intermediate (48) (Fig. 17.23), with a Kr (Equation 17.41) in the region of 10 M (123). Clearly, even a crude analog of (48) would have the potential to be an extremely powerful inhibitor of ADA. 

Leucine aminopeptidase is interesting in that its active site contains two zinc atoms which together bind and activate the water molecule [74]. Despite this enzyme containing a dinuclear metal center at its active site, its mechanism, and specifically its mode of proton transfers reactions, appear to follow the general theme established by thermolysin and carboxypeptidase Adenosine deaminase and other members of the family of nucleoside and nucleotide deaminases utilize zinc-bound water as the catalytic nucleophile to displace ammonia from the 6-position of purines or the 4-position of pyrimidines and in all cases display inverse solvent deuterium isotope effects ranging from 0.3 to 0.8 on fec/Kni [75-80]. These effects are reminiscent of those observed for metallopro-teases and have their origins, like those of the proteases, in fractionation factors for the protons of the bound water that are less than one. 

USA pegademase bovine enzyme Adagen Reaction product of recombinant adenosine deaminase with succinic anhydride and subsequent esterification with polyethylene glycol monomethyl ether Immuno- stimulant... 

Fig. 41.10. Salvage of bases. The purine bases hypoxanthine and gnanine react with PRPP to form the nucleotides inosine and gnanosine monophosphate, respectively. The enzyme that catalyzes the reaction is hypoxanthine-gnanine phosphoribosyltransferase (HGPRT). Adenine forms AMP in a reaction catalyzed by adenine phosphoribosyltransferase (APRT). Nucleotides are converted to nucleosides by 5 -nucleotidase. Free bases are generated from nncleosides by purine nucleoside phosphorylase. Deamination of the base adenine occurs with AMP and adenosine deaminase. Of the purines, only adenosine can be directly phosphorylated back to a nucleotide, by adenosine kinase.

Selective quenching allows also to put into evidence and to study a conformational change within a protein. ADAR2 (adenosine deaminase that acts on RNA) is a -80-kDa protein that efficiently deaminates the R/G site of GluR-B pre-mRNA sequences in vitro (O Connell et al. 1997). This enzyme has an RNA binding domain (RBD)l located in the C-terniinal catalytic domain. Deamination of adenosine (A) in the mRNA results in inosine (I) at that position. Because inosine is translated as guanosine (G), the editing reaction causes a functional A to G replacement. 

Pentostatin (deoxycoformycin) 4.18) and its ribose analogue, coformycin, strongly inhibit adenosine deaminase, the enzyme that converts adenosine 9.72) to inosine 9.74). presumably through the intermediate 9.73). This reaction is similar to the hydrolysis of an amidine to an amide, and it needs to be prevented when medicating patients with adenine-formulated drugs such as vidarabine 4.16). It can be seen that the structure of pentostatin resembles the... 

Pharma. Darmstadt, Germany) to give a stable eniyme preparation. Given the low solubility of (-) Carbovir, the concentration of (-)-2 was reduced to 2.5 g/iiter in bioconversions with immobilized enzyme so that the beads could easily be recovered from the reaction mixture without interf ence from the product. The enzyme was reused for up to 10 cycles without any significant loss of activity. This work demonstrated the potential of adenosine deaminase as a catalyst for large-scale production of optically pure (-)-cat-bovir. More recently, alternative routes to gain access to either enantiomer of carbovir have also been reported (40,41)-... 

---