Isomers, enzyme specificity

Enzymatic Method. L-Amino acids can be produced by the enzymatic hydrolysis of chemically synthesized DL-amino acids or derivatives such as esters, hydantoins, carbamates, amides, and acylates (24). The enzyme which hydrolyzes the L-isomer specifically has been found in microbial sources. The resulting L-amino acid is isolated through routine chemical or physical processes. The D-isomer which remains unchanged is racemized chemically or enzymatically and the process is recycled. Conversely, enzymes which act specifically on D-isomers have been found. Thus various D-amino acids have been... 

All amino acids except glycine exist in these two different isomeric forms but only the L isomers of the a-amino acids are found in proteins, although many D amino acids do occur naturally, for example in certain bacterial cell walls and polypeptide antibiotics. It is difficult to differentiate between the D and the L isomers by chemical methods and when it is necessary to resolve a racemic mixture, an isomer-specific enzyme provides a convenient way to degrade the unwanted isomer, leaving the other isomer intact. Similarly in a particular sample, one isomer may be determined in the presence of the other using an enzyme with a specificity for the isomer under investigation. The other isomer present will not act as a substrate for the enzyme and no enzymic activity will be demonstrated. The enzyme L-amino acid oxidase (EC 1.4.3.2), for example, is an enzyme that shows activity only with L amino acids and will not react with the D amino acids. 

It should be noted that some commercial enzyme preparations may contain several enzyme isomers (enzymes derived from one source which belong to the same enzyme class but differ in specificity, stability or other properties). This is most often the case when the commercial preparation was developed for a process industry application rather than a specific chemical biotransformation application. Some fungal enzymes, such as laccase, are sometimes supplied as crude enzyme mixtures. Fungal laccases are manufactured on a huge scale (multitonne per annum) and are principally used in bulk processes such as wood... 

It was first shown byE. Fischer, in 1894, that enzymes were specific in their action thus maltase acts only upon a-glucosides and emulsin only upon /3-glucosides. Later, he found that trypsin acted asymmetrically upon inactive polypeptides, e.g., alanyl-leucine was hydrolysed in such a way that only the compound composed of d-alanineand I-leucine, the natural isomers, was split up into its constituents, whereas the compound composed of 1-alanine and d-leucine was unattacked. Again, inactive leucine ester was found by Warbui to be only partially hydrolysed by trypsin he obtained 1-leucine and d-leucine ester. 

V-acetylmannosamine, but all at less than 5% of the natural substrate d-arabinose.58a This enzyme is specific for substrates with the -configuration at C(3), and Re-face addition of pyruvate to the aldehyde acceptors is the normal mode of attack.580 Though the enzymes accept substrates with C(2) hydroxyl of R- and -configurations, it prefers the S-isomer (Scheme 5.36). 

Enzymes are proteins which have the capability to catalyze many complex chemical reactions. Outstanding properties of these biological catalysts are their specificity and their capability of catalyzing the reaction of a substrate at very low concentration. Many enzymes are specific for a particular reaction of a particular substrate even in the presence of other isomers of that substrate or similar compounds. Some other enzymes are specific for a particular class of compounds. 

Studies of isomer-specific dephosphorylation by protein phosphatase 2a (PP2a) demonstrated that this enzyme is unable to dephosphorylate the ris-Serp/Thrp-Pro moieties in model peptides [55,56]. A similar specificity was reported for the pro-... 

There are two points to note carefully (1) Phosphorylation occurs specifically at C-6, the carbon furthest from the carbonyl group (Fig. 22-2), and (2) only the d carbohydrate isomers are involved in metabolism. In the future, we will omit the conformational identity from these equations for convenience only. It is critical to note that this reaction, like all others of metabolism, require a specific enzyme in order to proceed at a rate appropriate to the temperature and time requirements of metabolism. The enzyme is specific with respect to two issues (1) the transfer of a phosphate group (all enzymes specific for the transfer of a phosphate group are called kinases), and (2) the enzyme is selectively adapted to hexose sugars. 

The enzyme is specific for the 9-hydroperoxides and does not attack the 13-hydroperoxide isomers. The reaction mechanism is not known, but 0-labeling studies have shown that the ether oxygen is not derived from the —OOH group (Galliard and Matthew, 1975). 

The enzyme hydroxyacyl dehydrogenase described above is specific for the L-isomer. Apparently, some mammalian tissue can also oxidize the D-isomer, but it is not clear what enzymic mechanism is responsible for that reaction. Although the presence of an enzyme that specifically catalyzes the oxidation of the D-hy-droxyacyl ester to yield the keto acid has been proposed by some, others believe that the D-hydroxyacyl is transformed to the L-hydroxy acid by enzymes with racemase activity—namely, crotonase and another racemase. The equilibrium of the enzyme reaction is modified by the presence of magnesium in the medium. The modification of the equilibrium probably results from the complexion of magnesium with the keto acid. Eliminating the product favors the formation of the hydroxyacyl. Hydroxybutyrate can also be oxidized by an enzyme found in the mitochondria of many tissues, such as brain, kidney, heart, and liver. Hydroxybutyrate dehydrogenase has been isolated, solubilized, purified from beef heart, and demonstrated to require lecithin for activity. 

Just as previous studies showed, isomer selectivity was a common phenomenon in biodegradation of pyrethroids in soil, as observed for the permethrinase enzyme, which has a strong preference for the fran -isomer of permethrin. However, EstP, PytH and PytZ appear to have no isomer specificity (Maloney et al. 1993 Guo et al. 2009 Wu et al. 2006 Zhai et al. 2012 Russel et al. 2011). 

Step 2 of Figure 29.12 Isomerization Citrate, a prochiral tertiary alcohol, is next converted into its isomer, (2, 35)-isocitrate, a chiral secondary alcohol. The isomerization occurs in two steps, both of which are catalyzed by the same aconitase enzyme. The initial step is an ElcB dehydration of a /3-hydroxy acid to give cfs-aconitate, the same sort of reaction that occurs in step 9 of glycolysis (Figure 29.7). The second step is a conjugate nucleophilic addition of water to the C=C bond (Section 19.13). The dehydration of citrate takes place specifically on the pro-R arm—the one derived from oxaloacetate—rather than on the pro-S arm derived from acetyl CoA. 

Since endosulfan is a cytochrome P450-dependent monooxygenase inducer, the quantification of specific enzyme activities (e.g., aminopyrine-A -demethylase, aniline hydroxylase) may indicate that exposure to endosulfan has occurred (Agarwal et al. 1978). Because numerous chemicals and drugs found at hazardous waste sites and elsewhere also induce hepatic enzymes, these measurements are nonspecific and are not necessarily an indicator solely of endosulfan exposure. However, these enzyme levels can be useful indicators of exposure, together with the detection of endosulfan isomers or the sulfate metabolite in the tissues or excreta. 

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