Growth and translation elongation rate are sensitive to the concentration of EF‐Tu

Tubulekas, Ioannis; Hughes, Diarmaid

doi:10.1111/j.1365-2958.1993.tb01619.x

Cited by 39 publications

(36 citation statements)

References 62 publications

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“…1A). As the estimated Michaelis constant corresponds to the concentration at which translation would proceed at half of its maximal speed, these numbers indicate that the translation speed ranges between 70% and 86% of its maximum, consistent with earlier estimates for Salmonella (16). Our estimate of the Michaelis constant thus suggests that translation indeed operates close to the diffusion limit and that macromolecular crowding imposes a substantial cost to the cell by setting a large scale for the required allocation of T-proteins.…”

Section: Resultssupporting

confidence: 72%

See 1 more Smart Citation

Molecular crowding limits translation and cell growth

Klumpp

Scott

Pedersen

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

268

359

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Bacterial growth is crucially dependent on protein synthesis and thus on the cellular abundance of ribosomes and related proteins. Here, we show that the slow diffusion of the bulky tRNA complexes in the crowded cytoplasm imposes a physical limit on the speed of translation, which ultimately limits the rate of cell growth. To study the required allocation of ancillary translational proteins to alleviate the effect of molecular crowding, we develop a model for cell growth based on a coarse-grained partitioning of the proteome. We find that coregulation of ribosome-and tRNA-affiliated proteins is consistent with measured growth-rate dependencies and results in near-optimal allocation over a broad range of growth rates. The analysis further resolves a long-standing controversy in bacterial growth physiology concerning the growth-rate dependence of translation speed and serves as a caution against premature identification of phenomenological parameters with mechanistic processes. Bacterial cell growth and protein synthesis are tightly coupled as proteins account for a large fraction of the cellular biomass (1). In the model organism Escherichia coli, over half of the biomass is protein (2), and protein synthesis accounts for more than two-thirds of the cell's ATP budget during rapid growth (3). Therefore, the machinery of protein synthesis, i.e., ribosomes, tRNAs, and ribosome-affiliated factors, plays a central role in maintaining exponential growth (1, 4). This is manifested by an increased ribosome content in rapidly growing cells (2,5,6), by direct observations that protein synthesis is limited by the availability of free ribosomes (7), and by considerations that link evolutionary selective pressure to the cost of protein synthesis (8).The most striking evidence for the central role of ribosomes in cell growth is provided by the linear relation between the ribosome mass fraction and the growth rate for bacteria grown in media containing different nutrients. This linear relation, which emerged from the systematic characterization of bacterial cells growing at different rates (5, 9), is illustrated in Fig. 1A with data for E. coli (2, 10, 11). It can be interpreted as reflecting the intrinsically autocatalytic activity of ribosomes synthesizing ribosomal proteins (9, 12) and identifies the fraction of ribosomes allocated to making ribosomal proteins as a key determinant of the growth rate (11). The picture that emerges from such considerations has formed the basis of a systematic theory of bacterial growth, based on empirical "growth laws", similar to the phenomenological laws of physics (11,13). The theory provides a successful framework for the analysis of the interdependence of cell growth and gene expression, of the effects of antibiotics, and of protein overexpression (11) without the need to characterize how the individual steps of synthesis and degradation are affected by the global state of the cell (14).In addition to their high ribosome content, rapidly growing bacteria also contain large amounts of othe...

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Section: Resultssupporting

confidence: 72%

“…In particular, the most abundant protein in fastgrowing E. coli is elongation factor Tu (EF-Tu), with about six to seven molecules per ribosome (15), and accounting for up to 10% of the total protein mass under conditions of rapid growth (2,16). EF-Tu escorts charged tRNA to the ribosome and protects it from losing its amino-acylation (17,18).…”

mentioning

confidence: 99%

Molecular crowding limits translation and cell growth

Klumpp

Scott

Pedersen

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

268

359

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“…3d). This observation is in agreement with previous studies demonstrating the correlation between the cellular concentration of EF-Tu and organismal fitness (Brandis et al 2016;Tubulekas and Hughes 1993). Taken together, these results indicate that each experimental population exhibited parallel patterns of response such as upregulation of EF-Tu, as well as more idiosyncratic means of compensating for altered EF-Tu expression and activity.…”

Section: Promoter-level Mutations Upregulate Ancient Ef-tu Expressionsupporting

confidence: 82%

Experimental evolution of Escherichia coli harboring an ancient translation protein

Kaçar

Tran

et al. 2016

Preprint

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The ability to design synthetic genes and engineer biological systems at the genome scale opens new means by which to characterize phenotypic states and the responses of biological systems to perturbations. One emerging method involves inserting artificial genes into bacterial genomes, and examining how the genome and its new genes adapt to each other. Here we report the development and implementation of a modified approach to this method, in which phylogenetically inferred genes are inserted into a microbial genome, and laboratory evolution is then used to examine the adaptive potential of the resulting hybrid genome. Specifically, we engineered an approximately 700-million-year old inferred ancestral variant of tufB, an essential gene encoding Elongation Factor Tu, and inserted it in a modern Escherichia coli genome in place of the native tufB gene. While the ancient homolog was not lethal to the cell, it did cause a two-fold decrease in organismal fitness, mainly due to reduced protein dosage. We subsequently evolved replicate hybrid bacterial populations for 2,000 generations in the laboratory, and examined the adaptive response via fitness assays, whole-genome sequencing, proteomics and biochemical assays. We found that the hybrid lineages exhibit a general adaptive strategy in which the fitness cost of the ancient gene was ameliorated in part by up-regulation of protein production. We expect that this ancient-modern recombinant method may pave the way for the synthesis of organisms that exhibit ancient phenotypes, and that laboratory evolution of these organisms may prove useful in elucidating insights into historical adaptive processes.

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“…The choice of bacterial species and resistances used for these studies was based mainly on the availability of a genetically defined system with an established animal model rather than clinical importance. The streptomycin-resistant mutants have been characterized previously and classified into a restrictive and a nonrestrictive class with respect to their translation fidelity (12). One mutant from the nonrestrictive class and two from the restrictive class were chosen for further experiments.…”

Section: Resultsmentioning

confidence: 99%

Virulence of antibiotic-resistant Salmonella typhimurium

Björkman

Hughes

Andersson

1998

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

318

230

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We show that most Salmonella typhimurium mutants resistant to streptomycin, rifampicin, and nalidixic acid are avirulent in mice. Of seven resistant mutants examined, six were avirulent and one was similar to the wild type in competition experiments in mice. The avirulent-resistant mutants rapidly accumulated various types of compensatory mutations that restored virulence without concomitant loss of resistance. Such second-site compensatory mutations were more common then reversion to the sensitive wild type. We infer from these results that a reduction in the use of antibiotics might not result in the disappearance of the resistant bacteria already present in human and environmental reservoirs. Thus, second-site compensatory mutations could increase the fitness of resistant bacteria and allow them to persist and compete successfully with sensitive strains even in an antibiotic-free environment.During the last decade there has been an alarming increase in the appearance of antibiotic-resistant bacteria as a result of an increased use of antibiotics combined with the exceptional ability of bacteria to develop resistance. One strategy to reverse this development is to decrease the use of antibiotics to promote the disappearance of the resistant bacteria present in human and environmental reservoirs. Implicit in this reasoning is that resistance confers a cost on the bacteria, which results in a counter-selection against resistant strains in an antibiotic-free environment. An associated question and potential problem is whether the supposedly less fit, avirulentresistant bacteria might accumulate compensatory mutations that restore fitness and virulence without loss of resistance, and thereby stabilize the resistant population.In spite of the importance of these questions, there are few experiments that explicitly address them under the relevant conditions, i.e., in animal model systems using genetically defined bacterial strains (for a review, see ref. 1). For example, it has been shown in Escherichia coli that carriage of resistance genes on a plasmid is associated with a decreased growth rate, and that these strains can accumulate chromosomal compensatory mutations that, by an unknown mechanism, compensate for the growth rate decrease (2, 3). Likewise, it has been shown that slow-growing streptomycin-resistant mutants of E. coli can accumulate compensatory mutations that restore rapid growth under laboratory conditions without affecting the resistance (4). Interestingly, these compensatory mutations appear to create a genetic background in which the streptomycinsensitive revertants have a strong selective disadvantage, implying that it would be difficult for an evolved resistant strain to become sensitive even in the absence of the antibiotic (5). There are also animal data which indicate that tetracyclineresistant E. coli persist in pigs long after the antibiotic has been removed, suggesting that the added burden of this particular resistance in vivo is in fact small (6). Finally, studies of human clini...

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Growth and translation elongation rate are sensitive to the concentration of EF‐Tu

Cited by 39 publications

References 62 publications

Molecular crowding limits translation and cell growth

Molecular crowding limits translation and cell growth

Experimental evolution of Escherichia coli harboring an ancient translation protein

Virulence of antibiotic-resistant Salmonella typhimurium

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