Promiscuity catalytic

In principle, numerous reports have detailed the possibility to modify an enzyme to carry out a different type of reaction than that of its attributed function, and the possibility to modify the cofactor of the enzyme has been well explored [8,10]. Recently, the possibility to directly observe reactions, normally not catalyzed by an enzyme when choosing a modified substrate, has been reported under the concept of catalytic promiscuity [9], a phenomenon that is believed to be involved in the appearance of new enzyme functions during the course of evolution [23]. A recent example of catalytic promiscuity of possible interest for novel biotransformations concerns the discovery that mutation of the nucleophilic serine residue in the active site of Candida antarctica lipase B produces a mutant (SerlOSAla) capable of efficiently catalyzing the Michael addition of acetyl acetone to methyl vinyl ketone [24]. The oxyanion hole is believed to be complex and activate the carbonyl group of the electrophile, while the histidine nucleophile takes care of generating the acetyl acetonate anion by deprotonation of the carbon (Figure 3.5). 

This model clearly shows that the catalytic machinery involves a dyad of histidine and aspartate together with the oxyanion hole. Hence, it does not involve serine, which is the key amino acid in the hydrolytic activity of lipases, and, together with aspartate and histidine, constitutes the active site catalytic triad. This has been confirmed by constructing a mutant in which serine was replaced with alanine (Serl05Ala), and finding that it catalyzes the Michael additions even more efficiently than the wild-type enzyme (an example of induced catalytic promiscuity ) [105]. 

Finally, it should be mentioned that recently a new type of enzyme catalytic promiscuity reaction has been reported, which does not involve any of the catalytic... 

Decarboxylases are one of the members of the enolase superfamily. The most important and interesting point of this class of enzymes is that they are mechanistically diverse and catalyze different overall reactions. However, each enzyme shares a partial reaction in which an active site base abstracts a proton to form a nucleophile. The intermediates are directed to different products in the different active sites of different members. However, some enzymes of this class exhibit catalytic promiscuity in their natural form. ° This fact is considered to be strongly related to the evolution of enzymes. Reflecting the similarity of the essential step of the total reaction, there are some successful examples of artificial-directed evolution of these enzymes to catalyze distinctly different chemical transformation. The changing of decarboxylase to racemase described in Section 2.5 is also one of these examples. 

Kazlauskas, R.J., Enhancing catalytic promiscuity for biocatalysis. Curr. Opin. Chem. Biol.,... 

Old and new enzymes developed by this mechanism are members of the same superfamily. The main champions of this mechanism are Gerlt and Babbitt (Gerlt, 1999 Babbitt, 2000). Readers are also encouraged to consult the reference on catalytic promiscuity by O Brien and Herschlag (O Brien, 1999). 

In contrast to moonlighting proteins, which often are unknown to a researcher working in biocatalysis, proteins demonstrating catalytic promiscuity are fairly common and have also, in fact, been discussed in previous chapters of this book. Table 16.3 lists some examples of promiscuous enzymes, for a review, readers are referred to O Brien (1999). 

P. J. O Brien and D. Herschiag, Catalytic promiscuity and the evolution of new enzymatic activities, Chem. Bid. 1999, 6, R91-R105. 

Hatano, 1995b) from Amycolaptosis sp. is known to catalyze more than one different chemical reaction using a substantially different substrate (Palmer et al., 1999). Investigation of this catalytic flexibility in the context of the enolase superfamily raises the question of whether this enzyme may represent an example of nature s present-day reengineering of the superfamily scaffold for an entirely new function. Other examples of catalytically promiscuous enzymes from other superfamilies have been observed, as reviewed by O Brien and Herschlag (O Brien and Her-schlag, 1999). 

Enzyme catalytic promiscuity, where enzymes catalyze not only reactions with their natural substrates but also with non-natural substrates, has begun to be recognized as a valuable research and synthesis tool [6-10, 12, 13]. In particular, the catalytic promiscuity displayed by sucrose-utilizing transglucosidases toward a wide range of acceptor molecules has started to be exploited to provide novel synthesis pathways that are currently not available. 

Recently, even examples of lipase-catalyzed Michael additions and aldol condensations have appeared [7]. These are dramatic examples of catalytic promiscuity, that is, the ability of an enzyme to catalyze more distinctly different chemical transformations [8], Often such activities are explained in terms of the active site offering a scaffold in which substrates adopt favorable conformations and/ or reactants are brought together in a desired geometry. Accordingly, after being observed in wild-type enzymes, these side activities can often be enhanced in site-directed variants, in which residues in or close to the active site are mutated. 

Table 1 lists several examples for substrate ambiguity and catalytic promiscuity. The table indicates the EC numbers for the native and promiscuous activities, and thus categorizes their promiscuity accordingly to differences in EC numbers. Typical examples where differences in EC numbers reflect the degree of promiscuity, and cases where they might not are discussed below. 

Phosphotri-esterase from Phosphotriester hydrolysis EC Phosphodiesterase EC 3.1.4.X. Catalytic promiscuity 19, 86, 87... 

PLLs (PTE-like lactonases) Hydrolysis of quorum sensing Phosphotriester hydrolysis EC 3.1.8.1. Catalytic promiscuity 88... 

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