Cyclodextrin bis-imidazoles

Ribonuclease A is a member of a group of enzymes that cleave RNA using general acid-base catalysis without a metal ion in the enzyme. In ribonuclease A, such catalysis is performed by two imidazoles of histidine units, one as the free base (Im) and the other, protonated, as the acid (ImH+). To mimic this in an artificial enzyme, we prepared (3-cyclodextrin bis-imidazoles 41 [124]. The first one was a mixture of the... 

Cyclodextrin bis-imidazole catalyzes enolization by a bifunctional mechanism in which the ImH+ is hydrogen-bonded to the carbonyl oxygen while the Im removes the neighboring methyl proton (cf. 50). As expected from this, there was a bell-shaped pH vs. rate profile for the process. In the transition state two protons will move simultaneously, as in the hydrolysis reaction described above. Thus we indeed have a powerful tool to determine the geometric requirements for simultaneous bifunctional catalysis, a tool that could be of quite general use. 

The situation is complex. In another study we examined the cyclization of compound 54 catalyzed by cyclodextrin bis-imidazoles [140]. This dialdehyde can perform the intramolecular aldol reaction using the enol of either aldehyde to add to the other aldehyde, forming either 55 or 56. In solution with simple buffer catalysis both compounds are formed almost randomly, but with the A,B isomer 46 of the bis-imidazole cyclodextrin there was a 97 % preference for product 56. This is consistent with the previous findings that the catalyst promotes enolization near the bound phenyl ring, but in this case the cyclization is most selective with the A,B isomer 46, not the A,D that we saw previously. Again the enolization is reversible, and the selectivity reflects the addition of an enol to an aldehyde group. The predominant product is a mixture of two stereoisomers, 56A and 56B. Both were formed, and were racemic despite the chirality of the cyclodextrin ring. 

Figure 2.7 The AB Isomer of the cyclodextrin bis-imidazole Is the best catalyst for the hydrolysis of the catechol phosphate 9, indicating that the reaction proceeds through a phosphorane intermediate...

Figure 2.11 P-Cyclodextrin bis-imidazole is an acid/base catalyst for the enolization of p-tert-butylacetophenone, which binds into the cavity. The AD isomer is the most effective, indicating the preferred stereoelectronics of the enolization process...

FIGURE 12.2 Cyclodextrin bis(imidazoles) catalyzing hydrolysis of phosphate substrate. 

Two systems have been prepared so far. In one [4] we reacted the A,B isomer of cyclodextrin diiodide [5] with 6A,6B cyclodextrin bis-imidazole[5] (which is itself prepared from the same diiodide). The result was a mixture of two bis-imidazolium compounds, 2 which we call the "clam shell" type and the "love seat" type. 

With the evidence for our mechanism, it was of interest to construct a catalyst that could use it for cleavage of an RNA-like molecule. As we have described above, it is possible to attach imidazole rings to the primary carbons of cyclodextrins, and indeed we had prepared [20] two such catalysts that showed bifunctional catalysis of the hydrolytic cleavage of substrate M-However, we had designed our catalysts based on the previously accepted mechanism for RNA cleavage by ribonuclease. Since our mechanistic work revealed that a different mechanism was preferred, we redesigned the cyclodextrin bis-imidazole to place the imidazole rings in the correct position for the new mechanism. As hoped, this proved to be tiie best catalyst of all. 

A,6C and 6A,6D isomers (the seven glucose units of the cyclodextrin are labeled A through G). (As described below, we were later able to prepare as pure catalysts all the isomers of the bis-imidazole cyclodextrin with the imidazole rings on the primary carbons of the ring. The geometric dependence of catalysis indicated the mechanism involved.)... 

We examined the ability of our bis-imidazole cyclodextrin artificial enzymes to perform other bifunctionally-catalyzed reactions, where again the availability of the A,B and A,C and A.D isomers let us learn mechanistic details. As an important example, we examined three isomeric catalysts ability to promote the enolization of substrate 48, which binds into the cyclodextrin cavity in water [138]. Here there was again a strong preference among the isomers, but it was the A,D isomer 49 that was the effective catalyst It was also more effective than a cyclodextrin mono-imidazole that cannot use the bifunctional mechanism. 

Enolization can be part of an aldol condensation. We examined the aldol cyclization of compound 52 to 53 catalyzed by the bis-imidazole cyclodextrin artificial enzymes, and again saw that the A,D isomer was the preferred catalyst [139]. This was not an obvious result the rate-limiting step in this case is cyclization of the enol, which is... 

Tabushi, L Kuroda. Y. Bis(histamino)cyclodextrin-Zn-imidazole complex as an artificial carbonic anhydrase. J. Am. Chem. Soc. 1984,106 (16). 4580-4584. 

The pH vs rate profile showed a bell-shaped curve indicating that this catalyst uses both B and BH+ in a bifunctional mechanism. As with the enzyme, the bis-imidazole catalyst can perform its bifunctional catalysis by a simultaneous mechanism, not the sequential mechanism of simple buffer catalysis. We saw that this was indeed the case, as revealed by the tool called "proton inventory." In this technique the reaction is performed in D2O, in H2O, and in mixtures of the two. If only one proton that can exchange with D2O is moving in the transition state, the points all lie on a straight line between the H2O and slower D2O points. If two (or more) protons are moving, the line is curved. It had been found for the enzyme ribonuclease A [10] that a curved line was seen corresponding to the movement of two protons, and we also saw a curved plot—with very similar data— for our cyclodextrin-6A,6B-bisimidazole catalyst 6 [11]. Controls established that indeed this was a reliable indication that our system is performing simultaneous bifunctional catalysis, just as the enzyme does. In particular, the... 

Several model systems related to metalloenzymes such as carboxypeptidase and carbonic anhydrase have been reviewed. Breslow contributed a great deal to this field. He showed how to design precise geometries of bis- or trisimidazole derivatives as in natural enzymes. He was able to synthesize a modified cyclodextrin having both a catalytic metal ion moiety and a substrate binding cavity (26). Murakami prepared a novel macrocyclic bisimidazole compound which has also a substrate binding cavity and imidazole ligands for metal ion complexation. Yet the catalytic activities of these model systems are by no means enzymic. 

On treatment with potassium iodide, the capped disulfonate j8-cyclo-dextrin discussed above could easily be converted to the corresponding diiodide jS-cyclodextrin. With appropriate nucleophiles (imidazole, histamine) a new route to bis(iV-imidazolyl)-j8-cyclodextrin and bis(iV-histamino)-j8-cyclodextrin was developed by Tabushi s team (182). In the presence of Zn(II) ion, both regiospecifically bifunctionalized cyclodextrins hydrate CO2 and are the first successful carbonic anhydrase models. The Zn(II) ion binds to the imidazole rings located in the edge of the cyclodextrin pocket and the presence of an additional basic group, as with bis(histamino)-cyclodextrin-Zn(II), enhances the activity. Therefore, the present models show that all three factors, Zn(II)-imidazole, hydrophobic environment, and a base seem to help to generate the carbonic anhydrase activity (182). The chemistry of this enzyme is further discussed in Section 6.2, p. 331. 

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