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Journal of Bacteriology, February 2009, p. 1083-1091, Vol. 191, No. 3
0021-9193/09/$08.00+0     doi:10.1128/JB.00852-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Continued Protein Synthesis at Low [ATP] and [GTP] Enables Cell Adaptation during Energy Limitation{triangledown}

Michael C. Jewett,1,{dagger} Mark L. Miller,1 Yvonne Chen,1 and James R. Swartz1,2*

Department of Chemical Engineering,1 Department of Bioengineering, Stanford University, Stanford, California 943052

Received 22 June 2008/ Accepted 9 November 2008

One of biology's critical ironies is the need to adapt to periods of energy limitation by using the energy-intensive process of protein synthesis. Although previous work has identified the individual energy-requiring steps in protein synthesis, we still lack an understanding of the dependence of protein biosynthesis rates on [ATP] and [GTP]. Here, we used an integrated Escherichia coli cell-free platform that mimics the intracellular, energy-limited environment to show that protein synthesis rates are governed by simple Michaelis-Menten dependence on [ATP] and [GTP] (KmATP, 27 ± 4 µM; KmGTP, 14 ± 2 µM). Although the system-level GTP affinity agrees well with the individual affinities of the GTP-dependent translation factors, the system-level KmATP is unexpectedly low. Especially under starvation conditions, when energy sources are limited, cells need to replace catalysts that become inactive and to produce new catalysts in order to effectively adapt. Our results show how this crucial survival priority for synthesizing new proteins can be enforced after rapidly growing cells encounter energy limitation. A diminished energy supply can be rationed based on the relative ATP and GTP affinities, and, since these affinities for protein synthesis are high, the cells can adapt with substantial changes in protein composition. Furthermore, our work suggests that characterization of individual enzymes may not always predict the performance of multicomponent systems with complex interdependencies. We anticipate that cell-free studies in which complex metabolic systems are activated will be valuable tools for elucidating the behavior of such systems.

* Corresponding author. Mailing address: Department of Chemical Engineering, Stanford University, Stauffer III, Rm. 113, Stanford, CA 94305-5025. Phone: (650) 723-5398. Fax: (650) 725-0555. E-mail: jswartz{at}stanford.edu

{triangledown} Published ahead of print on 21 November 2008.

{dagger} Present address: Department of Genetics, Harvard Medical School, MA 02115.

Journal of Bacteriology, February 2009, p. 1083-1091, Vol. 191, No. 3
0021-9193/09/$08.00+0     doi:10.1128/JB.00852-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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