Enzymes isomerization

P-amylase, and debranching enzymes. Conversion of D-glucose to D-fmctose is mediated by glucose isomerase, mosdy in its immobilized form in columns. Enzymic degradation of starch to symps has been well reviewed (116—118), and enzymic isomerization, especially by immobilized glucose isomerase, has been fiiUy described (119) (see Syrups). 

To distinguish between simple, reversible slow binding (scheme B) and an enzyme isomerization mechanism (scheme C), one can examine the dependence of kobs on inhibitor concentration. If the slow onset of inhibition merely reflects inherently slow binding and/or dissociation, then the term kobs in Equations (6.1) and (6.2) will depend only on the association and dissociation rate constants k3 and k4 as follows ... 

For the enzyme isomerization mechanism illustrated in scheme C of Figure 6.3, there are two steps involved in formation of the final enzyme-inhibitor complex an initial encounter complex that forms under rapid equilibrium conditions and the slower subsequent isomerization of the enzyme leading to the high-affinity complex. The value of kohs for this mechanism is a saturable function of [/], conforming to the following equation ... 

In a two-step enzyme isomerization mechanism, as in scheme C, the affinity of the inhibitor encounter complex and the affinity of the final E I complex are reflected in the diminutions of v, and of vs, respectively, that result from increasing concen-... 

The form of Equation (6.7) reveals an interesting aspect of slow binding inhibiton due to enzyme isomerization. A slow forward isomerization rate is insufficient to result in slow binding behavior. The reverse isomerization rate must also be slow, and in fact must be significantly slower that the forward isomerization rate. If this were not the case, there would be no accumulation of the E I conformation at equilibrium. As the value of k6 becomes k5, the denominator of Equation (6.7) approaches unity. Hence the value of Kf approaches Kit and one therefore does not observe any time-dependent behavior. 

Both Vioxx and Celebrex derive their COX2 selectivity from the same type of isozyme-specific slow enzyme isomerization mechanism that was detailed here for DuP697. 

Lai, Z., et al., Human mdm2 mediates multiple mono-ubiquitination of p53 by a mechanism requiring enzyme isomerization. J Biol Chem, 2001, 276(33), 31357-67. 

A deficiency of vitamin A leads to vision defects, including a visual impairment at low light levels, termed night blindness. For the processes of vision, retinol needs to be converted first by oxidation into the aldehyde retinal, and then by enzymic isomerization to cw-retinal. c -Retinal is then bound to the protein opsin in the retina via an imine linkage (see Section 7.7.1) to give the red visual pigment rhodopsin. 

Slow transitions produced by enzyme isomerizations. This behavior can lead to a type of cooperativity that is generally associated with ligand-induced conformational changes . A number of enzymes are also known to undergo slow oligomerization reactions, and these enzymes may display unusual kinetic properties. If this is observed, it is advisable to determine the time course of enzyme activation or inactivation following enzyme dilution. See Cooperativity Bifurcation Theory Lag Time... 

Enzyme reaction mechanisms that contain one ligand-independent enzyme isomerization step. See Iso Mechanisms. 

A method used to assess enzyme mechanisms which may have one or more enzyme isomerization steps. In this procedure, [PJ/[P] is plotted as a function of time at different initial concentrations of substrate, where [PJ is the concentration of the product at time t and [Poo] is the concentration of P once the system has reached equilibrium. These data can be used to calculate the i iip term, associated with the isomerization step, in the rate expression. 

ENZYME EQUIVALENT WEIGHT ENZYME INHIBITION Enzyme isomerization,... 

At the present time, commercial isomerization processes based on enzymic catalysis are predominant, so only brief mention will be made of some of the nonenzymic processes that have been considered for commercialization in the past. Probably the major reasons for the current commercial use of enzymic rather than nonenzymic systems are that the nonenzymic systems so far developed result in products having one or more of the following defects too much ash, color, acid, off-flavor, a content of D-mannose or D-psicose, and high ratios of D-glucose to D-fructose. Probably, further advances in our understanding of the isomerization reaction and the mechanisms of the catalysis will lead to more efficient, nonenzymic processes that could replace the enzymic-isomerization systems now used commercially. 

Figure 3. Arrhenius plots for the rates of formation of intermediates in the reaction of papain with Na-carbobenzoxy-iL-lysine p-nitroanilide. The solvent was 60% dimethyl sulfoxide, pH 6.1, E0 = 3.0 X 10 6 M, = 3.0 X 10 5 M (32). Reactions 2 and 3 correspond to enzyme isomerization (32), Reaction 4 corresponds to the formation of the tetrahedral intermediate (32).

The fluorescent chlorophyll catabolites, such as pFCC (10), were observed not to accumulate during chlorophyll breakdown in senescent leaves (24). The indicated further transformation of the FCC chro-mophore to those of non-fluorescent chlorophyll catabolites (NCCs) was suggested to possibly be the result of a non-enzymic isomerization (56, 62). In analogy to the results of studies on the tautomerization chemistry of a range of hydro-porphinoids (91), the isomerization of the chromo-phore of FCCs into that of NCCs was judged to be rather favorable, thermodynamically. The complete de-conjugation of the four pyrrolic units, characteristic of the tetrapyrrolic NCCs, thus may occur in the course of natural chlorophyll breakdown under rather mild and, possibly, even without catalysis by (an) enzyme(s) (56). 

A less likely mechanism that produces oversaturation, but one which does apply to carbonic anhydrase, is one in which initial saturation with the substrate ties up enough enzyme for free enzyme isomerization to become rate limiting (i.e., E to E conversion limits V, rather than ES to E conversion) (46). Oversaturation then occurs at still higher substrate concentrations where there is more EA than free E, and EA conversion via E to E becomes rate limiting. With carbonic anhydrase, the basic reaction can be written... 

The countertransport or tracer perturbation method, introduced by Britton and Clarke (66), involves a test for obligate free enzyme isomerization (i.e., an Iso mechanism). The method was used by Knowles and co-workers to prove the Iso mechanism for proline racemase and show that V is limited by EA to E F, rather than by E to E, conversion (67). 

Two of the enzymes involved in the methionine cycle, namely, MTA nucleosidase [46] and MTR kinase [47] were purified from plants and characterised. However, the plant enzyme catalysing the formation of KMB from MTR-P has not been characterised. In rat liver extracts, three enzymes are involved in the conversion [42]. The first enzyme isomerizes MTR-P to l-phospho-5-methylthioribulose (MTRu-P), the second enzyme produces two unidentified metabolites from MTRu-P, and the third enzyme catalyses the conversion of these two metabolites to KMB with the uptake of oxygen. Plants may produce KMB from MTR-P by similar enzymes. The last step of the methionine cycle that converts KMB to methionine is most likely a transamination, and this activity was also detected in avocado extracts in the presence of asparagine [45]. 

Chiang LC, Hsiao HY, Ueng PP, Chen LF,Tsao GT (1982) Ethanol production from xylose by enzymic isomerization and yeast fermentation. In Scott CD (ed) Biotechnol Bioeng Symp. Wiley, New York, p 263... 

Some methods of metal ion-catalyzed chemical and enzymic isomerization (Lobry de Bruyn-Alberda van Ekenstein rearrangement, epimerization at C-2 of aldoses, action of isomerases) of free sugars have been reviewed (136 refs.). ... 

Enzyme isomerization reversible changes in enzyme conformation in the course of a catalytic cycle. See Enzyme kinetic parameters. 

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