Artificial enzymes approaches

In the enzyme design approach, as discussed in the first part of this chapter, one attempts to utilize the mechanistic understanding of chemical reactions and enzyme structure to create a new catalyst. This approach represents a largely academic research field aiming at fundamental understanding of biocatalysis. Indeed, the invention of functional artificial enzymes can be considered to be the ultimate test for any theory on enzyme mechanisms. Most artificial enzymes, to date, do not fulfill the conditions of catalytic efficiency and price per unit necessary for industrial applications. 

There is another approach that is increasingly part of synthesis the use of enzymes as catalysts. This approach is strengthened by the new ability of chemists and molecular biologists to modify enzymes and change their properties. There is also interest in the use of artificial enzymes for this purpose, either those that are enzyme-like but are not proteins, or those that are proteins but based on antibodies. Catalytic antibodies and nonprotein enzyme mimics have shown some of the attractive features of enzymes in processes for which natural enzymes are not suitable. 

In recent times the incorporation of enzymes into nanostructured materials is commonly referred to as nanobiocatalysis. Nanobiocatalysis has emerged as a rapidly growing research and development area. Lately, nanobiocatalytic approaches have evolved beyond simple enzyme immobilization strategies to include also topics like artificial enzymes and cells, nanofabrication, and nanopatterning [18]. A recent bibliometric analysis [19] of nanobiocatalysis publications shows a strong increase within the last decade (Fig. 14.1). The analysis has been compiled from... 

The seven chapters in this book describe various approaches to the synthesis and study of artificial enzymes. In Chapter 1,1 describe work in my laboratory over the past almost 50 years creating enzyme models and enzyme mimics. A major theme is the use of hydrophobic binding of substrates into cyclodextrins carrying catalytic groups,... 

It will be of some interest to learn how to build catalysts to handle the particular substrates that natural enzymes cleave, at a rate comparable to the rates of those enzymatic reactions. However, one of the aims of biomimetic chemistry is to extend the kinds of rates and selec-tivities of enzymatic reactions into reactions for which natural enzymes have not been optimized and to substrates that are neither recognized nor handled by normal enzymes. It is clear that we already have achieved this, even though our ribonuclease model system has some distance to go before it can approach the kinds of rates we have observed in the cyclodextrin ferrocinnamate ester reaction, for instance. In lock and key chemistry, the keys that fit artificial enzymes best are not the same as the keys that open the natural enzyme locks. 

Knor G. Bionic catalyst design a photochemical approach to artificial enzyme function. ChemBioChem 2001 2 593-6. 

This method of selecting catalytic sites significantly depends on spontaneous processes, in contrast to the development of artificial enzymes and catalytic antibodies. The selection process is based on self-assembly, selforganization and self-optimization. Therefore, this selection approach bears the characteristics of supramolecular chemistry. A similar concept is used in natural evolution processes, resulting in the complicated life forms we see around us today. Therefore, it is clear that we can design the self-organizational processes used in supramolecular chemistry to proceed according to the concepts followed by this natural evolutionary process. 

A much more improved antibody catalyst for amide hydrolysis has been elicited very recently by a joint hybridoma and combinatorial antibody library approach. The measured with a primary amide substrate at pH 9 and 25 °C was 5x10" s" for this new antibody. This corresponds to a half-life of 4 h when the substrate is fully complexed to the active site. The half-lives for the amide hydrolysis catalyzed by the antibodies are much longer than that (10-30 min at pH 4.5-7 and 4 °C when the substrate is fully complexed to the active site) of the light chain of Gbn hydrolyzed by 39. The fastest protein cleavage recorded so far with artiHcial proteinases is the cleavage of chymotrypsin by a coordinatively polymerized bilayer membrane which will be discussed later in this review. The coordinatively polymerized bilayer membrane achieved half-life as short as 3 min at 4 °C and pH 5.5-9.5. This is several times faster than the hydrolysis of Gbn by 39. In terms of utility in practical applications, however, 39 is more useful than the artificial enzyme based on the bilayer membrane due to the immobile nature of the former as well as the intrinsic instability of bilayer membranes. 

Catalytic antibodies, since the seminal contributions of Lerner and Schulz in the mid 1980s, have appeared as a fascinating way to construct artificial enzymes, specific for a given reaction. Some applications have appeared in asymmetric catalysis, for example Ref. [113]. However, it is too early to predict if this approach will develop into a useful methodology. 

Development of catalytic antibodies or abzymes is another fascinating approach to artificial enzymes. The concept of catalytic antibodies is that if a putative transition state analogue of a reaction for which a selective and efficient catalyst is desired is used to elicit antibodies. 

The catalytic power of abzymes is most conveniently measured as the rate acceleration compared to the uncatalyzed reaction. Although the early catalytic antibodies had only modest rate accelerations of a few hundred fold, more recent studies have produced artificial enzymes with catalytic activities almost approaching those of natural enzymes [576]. It must be noted, however, that in general, the rate accelerations of abzymes are in the range of 10 -10, though in rare cases they may reach values of 10. As yet, the corresponding values for most enzymes are far better (10 -10 ). Thus, in terms of catalytic efficiency, the most potent catalytic antibodies are just comparable to the slowest enzymes. 

Much more remains to be done with catalysts and reactions of the kind described in this paper. The selectivities achieved demonstrate that relatively available geometric factors can be used to control enzyme-like processes and achieve useful selectivities. However, one would expect that even more interesting systems could result if, for instance, we combined a coenzyme, a binding group, and a chirally placed catalytic group in transamination reactions. This would combine the two different approaches in the last two systems described. Work on the preparation of such compounds is underway, as is a general attempt to improve all of these systems so that artificial enzymes can play an increasing useful role in selective chemical synthesis [33]. 

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