Whole cells, of engineered

Whole cells of engineered E. coli BL21(DE3)(pMM4) desymmetrise cyclohexanones with substituents in the 4-position with variable results.36 Simple mono-substitution 118 and 120 gives good results but the enzyme is very sensitive to hydroxyl groups giving a lower yield with the tertiary alcohol 122 and poor ee with the secondary alcohol 124. 

Li L, Wang Y, Zhang L, Ma C, Wang A, Tao F, Xu P. (2012). Biocatalytic production of (2S,3S)-2,3-butanediol from diacetyl using whole cells of engineered Escherichia coli. Bioresour Technol, 115, 111-116. 

Engelking, H., Pfaller, R., Wich, G. and Weuster-Botz, D. (2006) Reaction engineering studies on /3-ketoester reductions with whole cells of recombinant Saccharomyces cerevisiae. Enzyme and Microbial Technology, 38, 536-544. 

Similarly, reductions of different racemic 2-oxabicyclo[3.2.0]heptan-6-ones [139], 2-oxabicyclo[2.2.1]heptane-7-carboxylates [140], norbomenone [102], and bicyclo[2.2.2]octan-2-one [141] have been performed the ee values, however, were moderate. For the reduction [141] of a 4-twistanone (tricyclo[4.4.0.0 ]decan-4-one), the alternative use of Rhodotorula rubra has been suggested. Kinetic resolution of racemic 5,6-epoxy-bicyclo[2.2.1]heptane-2-one using whole cells of genetically engineered S. cerevisiae allowed the synthesis of (+)-5,6-epoxy-bicyclo[2.2.1] heptane-2-ol [142]. 

Biocatalysts in nature tend to be optimized to perform best in aqueous environments, at neutral pH, temperatures below 40 °C, and at low osmotic pressure. These conditions are sometimes in conflict with the need of the chemist or process engineer to optimize a reaction with respect to space-time yield or high product concentration in order to facilitate downstream processing. Furthermore, enzymes and whole cells are often inhibited by products or substrates. This might be overcome by the use of continuously operated stirred tank reactors, fed-batch reactors, or reactors with in situ product removal [14, 15]. The addition of organic solvents to increase the solubility of substrates and/or products is a common practice [16]. 

Whole cells are grown for a variety of reasons. The cells may perform a desired transformation of the substrate, e.g., wastewater treatment the cells themselves may be the desired produce, e.g., yeast production or the cells may produce a desired product, e.g., penicillin. In the later case, the desired product may be excreted, as for the penicillin example, and recovered in relatively simple fashion. If the desired product is retained within the cell walls, it is necessary to lyse (rupture) the cells and recover the product from a complex mixture of cellular proteins. This approach is often needed for therapeutic proteins that are created by recombinant DNA technology. The resulting separation problem is one of the more challenging aspects of biochemical engineering. However, culture of the cells can be quite difficult experimentally and is even more demanding theoretically. 

A final problem for bioinformatics and bioanalytical scientists is the characterization of engineered microorganisms. Whole-cell analysis by mass spectrometry has been used to confirm the introduction of therapeutic genes into adenovirus vectors,100 to confirm the expression of recombinant proteins in bacteria,101,102 and also in vaccinology.103 In the broader case, identification of... 

Kaup, B., Bringer-Meyer, S. and Sahm, H. (2004) Metabolic engineering of Escherichia coir, construction of an efficient biocatalyst for D-mannitol formation in a whole-cell biotransformation. Applied Microbiology and Biotechnology, 64 (3), 333-339. 

During the last decade, significant advancements in biochemistry, molecular cloning, and random and site-directed mutagenesis, directed evolution of biocatalysts, metabolic engineering and fermentation technology have led us to devise methods to circumvent the disadvantages of whole-cell biotransformation discussed in Section 10.2. The applications of these methods are summarized in this section. 

As a perspective to this review, we conclude with recent developments in the use of whole cells to control the growth of nanopartides. Such approaches should allow one to take advantage of the whole cellular machinery and, combined with genetic engineering, appear very promising for the development of a green nanochemistry. 

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