Unit of Transcription in Bacteria The Operon

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19.1— Overview
To survive, a living cell must be able to respond to changes in its environment. One of many ways in which cells adjust to changes is to alter expression of specific genes, which, in turn, affects the number of corresponding protein molecules in the cell. This chapter focuses on some of the molecular mechanisms that determine when a given gene will be expressed and to what extent. The attempt to understand how expression of genes is regulated is one of the most active areas of biochemical research today.
It makes sense for a cell to vary the amount of a given gene product available under different conditions. For example, the bacterium Escherichia coli (E. coli) contains genes for about 3000 different proteins, but it does not need to synthesize all of these proteins at the same time. Therefore it regulates the number of molecules of these proteins that are made. The classic illustration of this phenomenon is the regulation of the number of b ­galactosidase molecules in the cell. This enzyme converts the disaccharide lactose into the monosaccharides, glucose and galactose. When E. coli is growing in a medium containing glucose as the carbon source, b ­
galactosidase is not required and only about five molecules of the enzyme are present in the cell. When lactose is the sole carbon source, however, 5000 or more molecules of b ­galactosidase occur in the cell. Clearly, the bacteria respond to the need to metabolize lactose by increasing the synthesis of b ­galactosidase molecules. If lactose is removed from the medium, the synthesis of this enzyme stops as rapidly as it began.
The complexity of eukaryotic cells means that they have even more extensive mechanisms of gene regulation than do prokaryotic cells. The differentiated cells of higher organisms have a much more complicated physical structure and often a more specialized biological function that is determined, again, by the expression of their genes. For example, insulin is synthesized in b cells of the pancreas and not in kidney cells even though the nuclei of all cells of the body contain the insulin genes. Molecular regulatory mechanisms facilitate the expression of insulin in pancreas and prevent its synthesis in kidney and other cells. In addition, during development of the organism appearance or disappearance of proteins in specific cell types is tightly controlled with respect to timing and sequence of developmental events.
As expected from the differences in complexities, far more is understood about the regulation of genes in prokaryotes than in eukaryotes. However, studies on the control of gene expression in prokaryotes often provide exciting new ideas that can be tested in eukaryotic systems. Sometimes, discoveries about eukaryotic gene structure and regulation alter the interpretation of data on the control of prokaryotic genes.
Several of the best studied examples of gene regulation in bacteria will be discussed, followed by some illustrations of the organization and regulation of related genes in the human genome. Finally, the use of recombinant DNA techniques to express some human genes of clinical interest will be presented.
19.2— Unit of Transcription in Bacteria:
The Operon
The single E. coli chromosome is a circular double­stranded DNA molecule of about four million base pairs. Most of the approximately 3000 E. coli genes are not distributed randomly throughout this DNA; instead, the genes that code for the enzymes of a specific metabolic pathway are clustered in one region of the DNA. In addition, genes for associated structural proteins, such as the 70 or so proteins that comprise the ribosome, are frequently adjacent to one another. Members of a set of clustered genes are usually coordinately regulated; they are transcribed together to form a "polycistronic" mRNA species that contains the coding sequences for several proteins. The term operon describes the
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complete regulatory unit of a set of clustered genes. An operon includes the adjacent structural genes that code for the related enzymes or associated proteins, a regulatory gene or genes that code for regulator protein(s), and control elements that are sites on the DNA near the structural genes at which regulator proteins act. Figure 19.1 shows a partial genetic map of the E. coli chromosome that gives locations of structural genes of some of the different operons.
When transcription of the structural genes of an operon increases in response to the presence of a specific substrate in the medium, the effect is known as induction. The increase in transcription of the b ­galactosidase gene when lactose is the sole carbon source is an example of induction. Bacteria also respond to nutritional changes by quickly turning off the synthesis of enzymes that are no longer needed. As will be described below, E. coli synthesizes the amino acid tryptophan as the end product of a specific biosynthetic pathway. However, if tryptophan is supplied in the medium, the bacteria do not need to make it themselves, and synthesis of enzymes for this metabolic pathway is stopped. This process is called repression. It permits the bacteria to avoid using their energy for making unnecessary and even harmful proteins.
Induction and repression are manifestations of the same phenomenon. In one case the bacterium changes its enzyme composition so that it can utilize a specific substrate in the medium; in the other it reduces the number of enzyme
Figure 19.1 Partial genetic map of E. coli. The locations of only a few of the genes identified and mapped in E. coli are shown here. Three operons discussed in this chapter are indicated. Reproduced with permission from Stent, G. S., and Calendar, R. Molecular Genetics, An Introductory Narrative. San Francisco: Freeman, 1978, p. 289; modified from Bachmann, B. J., Low, K. B., and Taylor, A. L. Bacteriol Rev. 40:116, 1976.