Regeneration Cofactors

Because enzymes can be intraceUularly associated with cell membranes, whole microbial cells, viable or nonviable, can be used to exploit the activity of one or more types of enzyme and cofactor regeneration, eg, alcohol production from sugar with yeast cells. Viable cells may be further stabilized by entrapment in aqueous gel beads or attached to the surface of spherical particles. Otherwise cells are usually homogenized and cross-linked with glutaraldehyde [111-30-8] to form an insoluble yet penetrable matrix. This is the method upon which the principal industrial appHcations of immobilized enzymes is based. 

Biocatalytic ledox reactions offer great synthetic utility to organic chemists. The majority of oxidase-catalyzed preparative bioconversions are still performed using a whole-ceU technique, despite the fact that the presence of more than one oxidoreductase in cells often leads to product degradation and lower selectivity. Fortunately, several efficient cofactor regeneration systems have been developed (160), making some cell-free enzymatic bioconversions economically feasible (161,162). 

Coexpression of Genes for Carbonyl Reductase and Cofactor-Regenerating Enzymes... 

Figure 8.15 Coexpression of genes for carbonyl reductase and cofactor-regenerating enzymes [llc,dj.

In some cases it is more attractive to use whole microbial cells, rather than isolated enzymes, as biocatalysts. This is the case in many oxidative biotransformations where cofactor regeneration is required and/or the enzyme has low stability outside the cell. By performing the reaction as a fermentation, i.e. with growing microbial cells, the cofactor is continuously regenerated from the energy source, e.g. glucose. 

Expensive stoichiometric cofactor need Cofactor regeneration ... 

Kragl, 1J., Kruse, W., Hummel, W. and Wandrey, C. (1996) Enzyme engineering aspects of biocatalysis cofactor regeneration as example. Biotechnology and Bioengineering, 52, 309-319. 

Wichmann, R. and Vasic-Racki, D. (2005) Cofactor regeneration at the lab scale. Advances in Biochemical Engineering/Biotechnology, 92, 225-260. 

Cofactor regeneration is a necessary prerequisite for an in-vitro application of oxidoreductase enzymes, as the cofactors are too expensive to be used in stoichiometric amounts (Fig. 43.2) [17, 18]. 

The principal strategies of cofactor regeneration - namely the enzymatic, chemical and electrochemical approach - are presented in Scheme 43.2 and have been reviewed recently [17, 21-23]. This chapter does not intend to be exhaustive rather, it focuses on the systems where a transition-metal complex and... 

Scheme 43.2 Principal strategies of cofactor regeneration (En production enzyme E2 regeneration enzyme).

Enzymatic cofactor regeneration can be subdivided into two categories the enzyme-coupled approach, where two different enzymes are used (one for the production reaction, and one for the regeneration reaction) and the substrate-coupled approach, where one and the same enzyme is used for both production and regeneration (E = E2). The most convenient and commonly used enzymatic regeneration systems are summarized in Table 43.1. 

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