E. coli is the most frequently used host for production of enzymes and other proteins by recombinant DNA technology. E. coli is preferable for its relative simplicity, inexpensive and fast high-density cultivation, well-known genetics, and large number of compatible molecular tools available. Despite all these advantages, expression and production of recombinant enzymes are not always successful and often result in insoluble and nonfunctional proteins. There are many factors that affect the success of cloning, expression, and mass production of enzymes by recombinant E. coli. In this paper, these critical factors and approaches to overcome these obstacles are summarized focusing controlled expression of target protein/enzyme in an unmodified form at industrial level. 1. Introduction In the past few years recombinant DNA technology has enabled scientists to produce a large number of diverse proteins, in microorganisms, that were previously unavailable, relatively expensive, or difficult to obtain in quantity [1]. While the expression of foreign genes has been reported in a variety of microorganisms and cell lines, most of this work utilizes E. coli for the cloning and expression of foreign genes [2]. Production of enzymes involves cloning of the appropriate gene into an expression vector under the control of an inducible promoter [3]. 2. Enzyme Production in E. coli The expression of recombinant proteins in cells in which they do not naturally occur is termed heterologous protein production. Bacterial expression systems are commonly used for production of heterologous gene products of both eukaryotic and prokaryotic origin [4]. The expression of heterologous proteins in E. coli, which is the bacterial system, is most widely and routinely used. A number of therapeutically important proteins are now produced as heterologous in E. coli. The first heterologous protein to be employed clinically was human insulin produced in E. coli, first approved in 1982, in UK, West Germany, The Netherland, and USA [5] (Table 1). Table 1: Some enzymes which are produced in bulk quantities and their industrial applications. 3. General Considerations of Selecting E. coli as Heterogeneous Protein Expression Host E. coli is widely used as the host for heterogeneous protein expression for the following advantages: (1) ease of growth and manipulation using simple laboratory equipment; (2) availability of dozens of vectors and host strains that have been developed for maximizing expression; (3) a wealth of knowledge about the genetics and physiology of E. coli; (4) expression can
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