Biocatalyst isolated enzyme processes

Immobilization is the process of adhering biocatalysts (isolated enzymes or whole cells) to a solid support. The solid support can be an organic or inorganic material, such as derivatized cellulose or glass, ceramics, metallic oxides, and a membrane. Immobilized biocatalysts offer several potential advantages over soluble biocatalysts, such as easier separation of the biocatalysts from the products, higher stability of the biocatalyst, and more flexible reactor configurations. In addition, there is no need for continuous replacement of the biocatalysts. As a result, immobilized biocatalysts are now employed in many biocatalytic processes. 

Here we will focus on the biochemical aspects. The techniques of isolating enzymes, the process of enzyme immobilisation and the behaviour of immobilised enzyme reactors are discussed in detail in the BIOTOL text Technological Applications of Biocatalysts", so will not deal with these aspects in detail here. In outline, however, once the desired enzyme is isolated, it is attached to a carrier material. In order to ascertain sufficient accessibility of the enzyme, a bifunctional spacer molecule is attached to the carrier ... 

The above two processes employ isolated enzymes - penicillin G acylase and thermolysin, respectively - and the key to their success was an efficient production of the enzyme. In the past this was often an insurmountable obstacle to commercialization, but the advent of recombinant DNA technology has changed this situation dramatically. Using this workhorse of modern biotechnology most enzymes can be expressed in a suitable microbial host, which enables their efficient production. As with chemical catalysts another key to success often is the development of a suitable immobilization method, which allows for efficient recovery and recycling of the biocatalyst. 

Hummel, W., Abokitse, K., Drauz, K. et al. (2003) Towards a large-scale asymmetric reduction process with isolated enzymes Expression of an (5)-alcohol dehydrogenase in E. coli and studies on the synthetic potential of this biocatalyst. Advanced Synthesis and Catalysis, 345 (1 + 2), 153-159. 

Compared with isolated enzymes, application of whole cells as biocatalysts is usually more economical since there is no protein purification process involved. Whole cells can be used directly in chemical processes, thereby greatly minimizing formulation costs. Whole cells are cheap to produce and no prior knowledge of genetic details is required. Microorganisms have adapted to the natural environment and produce both simple and complex metabolic products from their nutrient sources through complex, integrated pathways. 

The second level of decision making concerns the form of the biocatalyst, i.e., a whole cell, a cell organelle, an enzyme complex, a crude enzyme preparation, or an isolated enzyme, aU either free or immobilized. Availability, price, cofactor need, etc., are points that must be considered at this level. The costs associated with immobilization obviously should be regained by the possibility of developing a more efficient process. 

Abstract The use of various immobilized biocatalysts in industrial research and production will be introduced. The applied catalysts span the range from isolated enzymes to microbial whole cells, and even examples of the use of plant cells and mammalian cells could be found. Approximately 65 processes have been reviewed in this article, roughly 50% of which are actual production processes in the chemical industry. The remaining 50% refer to biocatalytic transformations which were carried out at laboratory scale up to pilot scale. In this review special attention was drawn to the range of transformable substrates and the variety of different supports. 

Space-time yield. The second metric is the space-time yield (STY) of the process (known also as productivity)(mass of product/reactor volume/time). The reactor size is determined by the STY, a higher value giving a smaller reactor (and hence capital cost). Reasonable values are in the region of 1 g/1 per hour for cells and around 10 g/1 per hour for isolated enzyme. Very often, with pharmaceutical processes it is necessary to fit a process into existing equipment, and hence the STY should be matched to the equipment size dependent upon the required production rate. In principle, by adding more biocatalyst the STY can be increased, provided there are no mixing or mass transfer limitations. 

For the aforementioned reasons the use of isolated enzymes is often preferred due to reduction in side reactions and higher productivities (see Ref. [12] for a review of this topic). However this brings other challenges such as the need for effective cofactor regeneration. The choice between enzyme and whole-cell biocatalysts is complex and requires more work in the future to establish a clearer strategy to help the process design and implementation of bioreductions. 

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