Whole cells disadvantage

Biotransformations are carried out by either whole cells (microbial, plant, or animal) or by isolated enzymes. Both methods have advantages and disadvantages. In general, multistep transformations, such as hydroxylations of steroids, or the synthesis of amino acids, riboflavin, vitamins, and alkaloids that require the presence of several enzymes and cofactors are carried out by whole cells. Simple one- or two-step transformations, on the other hand, are usually carried out by isolated enzymes. Compared to fermentations, enzymatic reactions have a number of advantages including simple instmmentation reduced side reactions, easy control, and product isolation. 

Such isolated enzyme approaches for deracemization have a clear disadvantage in that they require two operational manipulations with an intermediate recovery step. A one-pot strategy is offered by employing whole-cell biotransformations with strains containing set(s) of complementary dehydrogenases operating in both biooxidative and bioreductive modes. Trace amounts of the intermediate ketone species can be isolated in several cases. In order to lead to an efficient deracemization... 

Prokaryotic cells express hundreds to thousands of proteins while higher eukaryotes express thousands to tens of thousands of proteins at any given time. If these proteins are to be individually identified and characterized, they must be efficiently fractionated. One-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) has typically been use to study protein mixtures of <100 proteins. Onedimensional electrophoresis is useful because nearly all proteins are soluble in SDS, molecules ranging from approximately 10,000 to 300,000 molecular weight can be resolved, and extremely basic or acidic proteins can be visualized. The major disadvantage to one-dimensional gels is that they are not suitable for complex mixtures such as proteins from whole cell lysates. 

Disadvantages of Whole-Cell Process Compared with the Isolated... 

Approaches to Address the Disadvantages of Whole-Cell Biotransformation... 

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. 

One of the major disadvantages of utilizing enzymes or semisynthetic enzymes for chemical transformations is the fact that large quantities of pure enzyme are needed for preparative scale. This disadvantage is contrasted with whole cell systems (bacteria, fungi, plant/animal cells) because they are easily available in large quantities through... 

Enzymic asynmietric epoxidation of alkenes may be performed by pure monooxygenases. However, due to practical problems such as need of cofactors, microbial oxidation with whole cells has been more widely used for the purpose. One great disadvantage however, is the toxicity of epoxides towards living cells. 

Mention some of the advantages and disadvantages of using isolated enzymes rather than whole cells in biocatalysis. Is it always needed to purify enzymes for a biocatalytic application To what degree should one purify ... 

In vitro systems with varying levels of complexity have been used to study xenobiotic metabolism. These include purified proteins, subcellular fractions/cell lysates, intact whole cells, tissue slices and whole organ culture. Advantages and disadvantages of the approaches are summarized below. 

Isolated enzymes are advantageous over whole cells owing to the absence of side activities compromising selectivity and the greater ease of repeated re-use to achieve high total turnover numbers (TTNs). Principal disadvantages are the requirement for a purification protocol, the yield loss upon purification, and the requirement for external addition of cofactors. 

In general there are two principle possibilities using a biocatalyst in organic synthesis, namely as whole cells or as isolated enzymes - free or immobilized. The advantages and disadvantages of each can be intensively discussed, but the outcome of this consideration always depends on the whole system and the kind of application. There are numerous examples of both and thus there is no partitioning between whole cell biotransformation and isolated enzymes in this review. 

Describe in your own words the advantages and disadvantages of using whole cells compared to isolated enzymes as biocatalysts, with respect to catalyst immobilization, catalyst recovery, ease of use, and product selectivity and purification. 

A disadvantage of this type of technique is that the impedance of the whole cell is measured, whereas in the investigation of electrode processes one is interested in the properties of one of the electrodes. It is possible to reduce the contribution of the unwanted components by using an auxiliary electrode with an area large relative to that of the electrode being studied, and extrapolating the cell impedance to infinite frequency in order to remove contributions such as cell resistance. 

There are different possibilities for performing whole cell biotransformations with several enzymes. Each enzyme can be produced from another strain. To carry out the desired reaction, these strains are combined in different amounts which depend on the particular enzyme activities. Disadvantages of this method are diffusion problems and the membrane barrier [126]. Alternatively, all enzymes can be expressed in one single host strain. This can be reached by means of recombinant DNA techniques in several ways The genes can be expressed under one single promoter or under different promoters. Furthermore, it is possible to express all genes from the same promoter but with different copy numbers. Using these methods it is possible to create tailor-made catalysts for manifold purposes. 

---