0-Galactosidase induction

Escherichia coli BR513 X-lac fusion, envA, uvrB 6-Galactosidase induction... 

Figure 31.2. P-Galactosidase Induction. The addition of lactose to an E. coli culture causes the production of P-galactosidase to increase from very low amounts to much larger amounts. The increase in the amount of enzyme parallels the increase in the number of cells in the growing culture. P-Galactosidase constitutes 6.6% of the total protein synthesized in the presence of lactose.

FIGURE 31.1 Kinetics of p-galactosidase induction by lactose and IPTG. (Assume that each inducer has been removed after an appropriate period.)... 

The kinetics of galactosidase induction are simple [102,103] on addition of excess inducer to growing cells, enzyme activity increases at a rate proportional to the increase in total protein within the culture. Enzyme formation reaches its maximum rate at 37°C within 3 minutes of addition of inducer. After removal of inducer, enzyme synthesis ceases in about the same amount of time. 

The characterization of the nar promoter in E. coli with the intact nar operon on the chromosome was carried out. Expression of P-galactosidase was maximal when the nar promoter was induced at ODgoo =1-7 under anaerobic conditions in the presence of 1 % nitrate. At this optimal condition the induction ratio was approximately 250 and the specific P-galactosidase activity was approximately 7500 Miller units at ODgoo = 2.7 [35]. In this study, we used plasmid... 

The maximum specific (3-galactosidase activities were also strongly dependent on the DO levels before and after induction. Maintenance of high DO levels before induction and low DO levels during the induction period was important to maximize the expression of (3-galactosidase (Table 2, rows C, E, F, and G). 

In the absence of nitrate, the maximum specific (3-galactosidase activity became only 4000 Miller units (Table 2, row H). Therefore, addition of nitrate into the growth medium before induction is necessary to maximize the expression of (3-galactosidase. 

Of three cases of different cell densities (ODgoo = 18, 35, 80) at the induction time, the highest value of specific P-galactosidase activity was obtained when... 

Hamilton, I. R. and Lo, G. C. 1978. Co-induction of 0-galactosidase and the lactose-phosphoenolpyruvate phosphotransferase system in Streptococcus salivarius and Streptococcus mutans. J. Bacteriol 136, 900-908. 

In the induction of enzymes of galactose metabolism in E. coli, three enzymes are involved -galactosidase (which catalyses the hydrolysis of the y -glycosidic bonds of lactose), galactose permease (which is responsible for transport of lactose across the cell membrane) and a third enzyme, A-protein, apparently not directly involved in galactose metabolism. The system has an environmental inducer, galactose, and in its presence the number of /)-galactosidase molecules rises from 5-10 to 10,000 within the cell. The addition of the inducer can increase the protein production in less than five minutes after its addition. Protein synthesis of these enzymes stops almost immediately in the absence of lactose. 

Peluso, R., Godwin, S., Carter, B. J., Gregory, R. J., Wadsworth, S. C. and Cheng, S. H. (2004). AAV2 vector harboring a liver-restricted promoter facilitates sustained expression of therapeutic levels of alpha-galactosidase A and the induction of immune tolerance in Fabry mice. Mol. Ther. 9, 231-240. 

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