Biochemical Engineering By D G Rao Free Download 12
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Although the capabilities of cell-free systems have increased over the years, researchers are working to further improve the systems to increase their capabilities. One of the biggest improvements to the cell-free system over the past three decades has been the introduction of continuous-exchange formats that allow cell-free systems to be maintained in a continuous, non-batch, and bioreactor-like manner. Utilizing these formats, researchers have been able to continuously provide amino acids, nucleotides, and energy sources in order to maintain a pool of reaction components while facilitating protein synthesis. Further, researchers have been able to add features that improve protein synthesis by maintaining concentration gradients of reaction components across dialysis membranes . In addition to continuous-exchange formats, researchers have been able to expand the types of lysates that can be used to prepare cell-free systems. Previously, early studies were limited to autolytic lysates with limited metabolic capacity . Adding a supplemental metabolic system to a cell-free system was a critical step in improving protein production. Calhoun et al. discovered that by supplementing a cell-free system with an addition of a fully functioning, intact bacterial cell, it was possible to produce far more than the original cell-free system with the addition of exogenous metabolic substrates . The addition of an intact cell allowed for the continuous replenishment of intracellular reactions, which is an essential feature of a cell-free system.
Additionally, cell-free systems are also being applied to problems in biotechnology and medicine through the optimization of enzymatic reaction parameters. Taylor et al. demonstrated that by changing reaction conditions and variables such as the concentration of lysate and substrate, it is possible to increase protein yield as well as reaction duration. These researchers also demonstrated that the ratio of amino acids to mRNA transcript can be altered to improve the yield of the protein . A similar method of improving cell-free synthesis was demonstrated by the Harvey Lab at the University of California, Riverside by varying the addition of amino acids, nucleotides, and enzyme concentrations to improve yields. This work also demonstrated that high concentrations of reaction components can have a negative impact on protein yield .
 Fagerlund, J. M.; Garcia-Lozano, J. E.; Ogden, D. N.; & Lieber, C. M. Determination of factors that limit protein synthesis in a cell-free system. Proceedings of the National Academy of Sciences of the United States of America. 1990;87(5):1703-1707.[/author_list] 827ec27edc