Engineering enzymatic catalysis

Wells, T.N.C., Fersht, A.R. Hydrogen bonding in enzymatic catalysis analyzed by protein engineering. Nature 316 656-657, 1985. 

Because practically all aromatic organic pollutants that release phenols or anilines in the course of their degradation could bind HS through enzymatic catalysis, methods employing enzyme-catalyzed polymerization reactions minimizing their presence by partial removal in aquatic and terrestrial environments might be utilized in pollution control. This can have a remarkable effect in environmental engineering practice. 

Colin J. Jackson was born in Hamilton (New Zealand) and received his B.Sc. (Hons) in 2002 from the University of Otago. In 2006 he completed his Ph.D. under the supervision of Professor David Ollis at the Australian National University. He subsequently completed a short postdoctoral fellowship under Dr. John Oakeshott at the Commonwealth Scientific and Industrial Research Organization (CSIRO, Australia) before being appointed as a research scientist. He is currently on leave to take up a Marie Curie Research Fellowship at the Institut de Biologie Structurale in Grenoble, France. He is broadly interested in the structural and chemical determinants of enzymatic catalysis and enzyme engineering. 

Enzymes have high potential in organic synthesis. Applications have been until now mainly based on kinetic resolution, but many opportunities exist to use enzymatic catalysis for enantioselective syntheses. Recently, it was shown by Reetz et al. that a combination of genetic engineering and mutagenesis can easily provide modified enzymes of greatly improved stereoselectivity for the transformation of a given substrate [114]. This concept should find wide applications in catalyzed enantioselective reactions. 

On the basis of past interest in the field, we have hopes that our conclusions will be of value not only to physical chemists concerned with the understanding of structure-reactivity problems, but to the industrial chemist and chemical engineer who desires a higher predictability for acid catalysed systems and to the biochemist who is concerned with proton transfer properties in enzymatic catalysis. 

With at least partial understanding of essential features of enzymatic catalysis has come the desire to improve, by the modification of existing enzymes through genetic or chemical means to alter their substrate specificity without loss of catalytic efficiency, the field of protein engineering. A unique feature in the antibody design is that the stereospecific catalysis can be sought for reactions not known to be enzyme catalyzed. 

Lipases are able to work in very different media. They work in biphasic systems and in monophasic (in the presence of hydrophilic or hydrophobic solvents) systems where the water content can vary significantly between aqueous and anhydrous media. They have been tested also in ionic liquid media (Lau et al. 2000 Wasserscheid and Keim 2000 Kamal and Chouhan 2004 Ha et al. 2007), in supercritical fluids (Laudani et al. 2007) and in gaseous media (Cameron et al. 2002). The different media for enzymatic catalysis has been outlined before (see section 1.6) and it will not be further discussed here. However, some examples of modulation of activity and selectivity of lipases by medium engineering will be described in this section. 

Weatherley, L.R. and Rooney, D. (2007). Enzymatic catalysis and electrostatic process intensification for processing of natural oUs. Chemical Engineering Journal, (In Press). 

The impact of biocatalysis in the fats and oils industry has not been particularly overwhelming. However, the causative factors for this are slowly being unravelled. It seems that the marriage between protein engineering, enzyme technology and conventional bioengineering will eventually be a sweet one. Enzymatic catalysis is expected to be the power house of the biochemical industries of tomorrow. 

Kobayashi S, Ohmae M (2(X)7) Polymer synthesis and modification by enzymatic catalysis. In Matyjaszewski K, Gnanou Y, Leibler L (eds) Macromolecular engineering precise synthesis, materials properties, applications, vol 10, Wiley-VCH, Weinheim., pp 400-477... 

If an appropriate selection can be made from the assorted mutations that block each step in enzymatic catalysis and lead to the accumulation of intermediates, such mutant enzymes can serve as direct tools to delineate a complex catalytic pathway. From both theoretical and practical standpoints, mutations and mutagenesis have significant impacts on genetic engineering. Generating mutant enzymes (or proteins) with either improved catalytic (or functional) and/or structural features or de novo specificities is a lofty goal of protein engineering. 

What can we learn about mechanism from protein engineering that cannot be learned from classical enzymology Chapter 7 begins with the statement The mechanism of an enzymatic reaction is ultimately defined when all the intermediates, complexes, and conformational states of an enzyme are characterized and the rate constants for their interconversion are determined. The classical delineation of a mechanism would have been achieved when the general nature of intermediates on a pathway and the type of catalysis had been determined. But... 

Further advantages of biocatalysis over chemical catalysis include shorter synthesis routes and milder reaction conditions. Enzymatic reactions are not confined to in vivo systems - many enzymes are also available as isolated compounds which catalyze reactions in water and even in organic solvents [28]. Despite these advantages, the activity and stability of most wild-type enzymes do not meet the demands of industrial processes. Fortunately, modern protein engineering methods can be used to change enzyme properties and optimize desired characteristics. In Chapter 5 we will outline these optimization methods, including site-directed mutagenesis and directed evolution. 

Hydrophilicity is an important criterion for the use of synthetic polymers. Existing methods for surface modihcation of synthetic hbers are costly and complex. Therefore, the enzymatic surface modihcation of synthetic hbers is a new and green approach to synthesize polymers with improved surface properties. Use of enzymes for surface modihcation of polymers will not only minimize the use of hazardous chemicals but also minimize the environment pollution load. Besides these, the enzyme-modihed polymers can also immobilize those enzymes which can only bind to the selective functional groups present on the polymeric surface such as —COOH and —NH2. Similarly, substrates can immobilize on the solid matrix (or polymer), which will be easily accessible to the enzymes. Genetic engineering can be employed for the modihcation of active sites of enzymes for better polymer catalysis. 

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