A computational model to uncover the biophysical underpinnings of neural firing heterogeneity in dissociated hippocampal cultures.

Calcium (Ca2+ ) imaging reveals a variety of correlated firing in cultures of dissociated hippocampal neurons, pinpointing the non-synaptic paracrine release of glutamate as a possible mediator for such firing patterns, although the biophysical underpinnings remain unknown. An intriguing possibility is that extracellular glutamate could bind metabotropic receptors linked with inositol trisphosphate (IP3 ) mediated release of Ca2+ from the endoplasmic reticulum of individual neurons, thereby modulating neural activity in combination with sarco/endoplasmic reticulum Ca2+ transport ATPase (SERCA) and voltage-gated Ca2+ channels (VGCC). However, the possibility that such release may occur in different neuronal compartments and can be inherently stochastic poses challenges in the characterization of such interplay between various Ca2+ channels. Here we deploy biophysical modeling in association with Monte Carlo parameter sampling to characterize such interplay and successfully predict experimentally observed Ca2+ patterns. The results show that the neurotransmitter level at the plasma membrane is the extrinsic source of heterogeneity in somatic Ca2+ transients. Our analysis, in particular, identifies the origin of such heterogeneity to an intrinsic differentiation of hippocampal neurons in terms of multiple cellular properties pertaining to intracellular Ca2+ signaling, such as VGCC, IP3 receptor, and SERCA expression. In the future, the biophysical model and parameter estimation approach used in this study can be upgraded to predict the response of a system of interconnected neurons.

Mutants lacking global regulators, fis and arcA, in Escherichia coli enhanced growth fitness under acetate metabolism by pathway reprogramming.

Global regulatory transcription factors play a significant role in controlling microbial metabolism under genetic and environmental perturbations. A system-level effect of carbon sources such as acetate on microbial metabolism under disrupted global regulators has not been well established. Acetate is one of the major substrates available in various nutrient niches such as the mammalian gut and a keto diet. A substantial amount of acetate gets secreted in aerobic metabolism. Therefore, investigating the study on acetate metabolism is highly significant. It is known that the global regulators fis and arcA regulate acetate uptake genes in E. coli under glucose conditions. This study deciphered the growth and flux distribution of E. coli transcription regulatory knockouts Δfis, ΔarcA and double deletion mutant, ΔarcAΔfis under acetate using 13C-metabolic flux analysis (MFA), which has not been investigated before. We observed that the mutants exhibited an expeditious growth rate (~ 1.2-1.6-fold) with a proportionate increase in acetate uptake rates compared to the wild type. 13C-MFA displayed the distinct metabolic reprogramming of intracellular fluxes via the TCA cycle, anaplerotic pathway and gluconeogenesis, which conferred an advantage of a faster growth rate with better carbon usage in all the mutants. This resulted in higher metabolic fluxes through the TCA cycle (~ 18-90%), lower gluconeogenesis (~ 15-35%) and higher CO2 and ATP production with the proportional increase in growth rate. The study reveals a novel insight by stating the sub-optimality of the wild-type strain grown under acetate substrate aerobically. These mutant strains efficiently oxidize acetate, thus acting as potential candidates for the biosynthesis of isoprenoids, biofuels, vitamins and various pharmaceutical products.Key Points• Mutants exhibited a better balance between energy and precursor synthesis than WT.• Leveraged in the unravelling of regulatory control under various nutrient shifts.• Metabolic readjustment resulted in optimal biomass requirement and faster growth.

Exploring the druggable proteome of Candida species through comprehensive computational analysis.

Candida albicans and non-albicans Candida spp. are major cause of systemic mycoses. Antifungal drugs such as azoles and polyenes are not efficient to successfully eradicate Candida infection owing to their fungistatic nature or low bioavailability. Here, we have adopted a comprehensive computational workflow for identification, prioritization and validation of targets from proteomes of Candida albicans and Candida tropicalis. The protocol involves identification of essential drug-target candidates using subtractive genomics, protein-protein interaction network properties and systems biology based methods. The essentiality of the novel metabolic and non-metabolic targets was established by performing in silico gene knockouts, under aerobic as well as anaerobic conditions, and in vitro drug inhibition assays respectively. Deletion of twelve genes that are involved in amino acid, secondary metabolite, and carbon metabolism showed zero growth in metabolic model under simulated conditions. The algorithm, used in this study, can be downloaded from http://pbit.bicnirrh.res.in/offline.php and executed locally.

Investigating host-bacterial interactions among enteric pathogens.

BACKGROUND: In 2017, World Health Organization (WHO) published a catalogue of 12 families of antibiotic-resistant "priority pathogens" that are posing the greatest threats to human health. Six of these dreaded pathogens are known to infect the human gastrointestinal system. In addition to causing gastrointestinal and systemic infections, these pathogens can also affect the composition of other microbes constituting the healthy gut microbiome. Such aberrations in gut microbiome can significantly affect human physiology and immunity. Identifying the virulence mechanisms of these enteric pathogens are likely to help in developing newer therapeutic strategies to counter them. RESULTS: Using our previously published in silico approach, we have evaluated (and compared) Host-Pathogen Protein-Protein Interaction (HPI) profiles of four groups of enteric pathogens, namely, different species of Escherichia, Shigella, Salmonella and Vibrio. Results indicate that in spite of genus/ species specific variations, most enteric pathogens possess a common repertoire of HPIs. This core set of HPIs are probably responsible for the survival of these pathogen in the harsh nutrient-limiting environment within the gut. Certain genus/ species specific HPIs were also observed. CONSLUSIONS: The identified bacterial proteins involved in the core set of HPIs are expected to be helpful in understanding the pathogenesis of these dreaded gut pathogens in greater detail. Possible role of genus/ species specific variations in the HPI profiles in the virulence of these pathogens are also discussed. The obtained results are likely to provide an opportunity for development of novel therapeutic strategies against the most dreaded gut pathogens.

Effect of global transcriptional regulators on kinetic behavior of Escherichia coli under anaerobic fermentation conditions.

The phenotype of Escherichia coli is governed by global transcriptional regulators under variable environmental conditions. Fnr, ArcA, IhfA-B, Crp, and Fis are amongst the major global transcription regulators that change their activity across the range of aeration, hence forming the core transcriptional network responsible for survival under changing aeration conditions. Effect of deletion of these global transcription factors on the kinetics of cell growth and mixed acid production under anaerobic fermentation conditions has not been characterized. To quantify the kinetic parameters in the absence of global transcription factors, experiments were performed using single deletion mutants of the above-mentioned global transcription regulators. The absence of global transcription regulators resulted in a relatively higher glucose uptake rate than that required for the observed growth rate. This further resulted in a higher yield of mixed acids per unit biomass in mutants as compared to the parent strain (E. coli BW25113). Thus, the increased channeling of carbon towards mixed acid secretion resulted in a lower growth rate in the mutants.

Role of phosphate limitation and pyruvate decarboxylase in rewiring of the metabolic network for increasing flux towards isoprenoid pathway in a TATA binding protein mutant of Saccharomyces cerevisiae.

BACKGROUND: Production of isoprenoids, a large and diverse class of commercially important chemicals, can be achieved through engineering metabolism in microorganisms. Several attempts have been made to reroute metabolic flux towards isoprenoid pathway in yeast. Most approaches have focused on the core isoprenoid pathway as well as on meeting the increased precursors and cofactor requirements. To identify unexplored genetic targets that positively influence the isoprenoid pathway activity, a carotenoid based genetic screen was previously developed and three novel mutants of a global TATA binding protein SPT15 was isolated for heightened isoprenoid flux in Saccharomyces cerevisiae. RESULTS: In this study, we investigated how one of the three spt15 mutants, spt15_Ala101Thr, was leading to enhanced isoprenoid pathway flux in S. cerevisiae. Metabolic flux analysis of the spt15_Ala101Thr mutant initially revealed a rerouting of the central carbon metabolism for the production of the precursor acetyl-CoA through activation of pyruvate-acetaldehyde-acetate cycle in the cytoplasm due to high flux in the reaction caused by pyruvate decarboxylase (PDC). This led to alternate routes of cytosolic NADPH generation, increased mitochondrial ATP production and phosphate demand in the mutant strain. Comparison of the transcriptomics of the spt15_Ala101Thr mutant cell with SPT15WT bearing cells shows upregulation of phosphate mobilization genes and pyruvate decarboxylase 6 (PDC6). Increasing the extracellular phosphate led to an increase in the growth rate and biomass but diverted flux away from the isoprenoid pathway. PDC6 is also shown to play a critical role in isoprenoid pathway flux under phosphate limitation conditions. CONCLUSION: The study not only proposes a probable mechanism as to how a spt15_Ala101Thr mutant (a global TATA binding protein mutant) could increase flux towards the isoprenoid pathway, but also PDC as a new route of metabolic manipulation for increasing the isoprenoid flux in yeast.

Enhancement of Swimming Speed Leads to a More-Efficient Chemotactic Response to Repellent.

Negative chemotaxis refers to the motion of microorganisms away from regions with high concentrations of chemorepellents. In this study, we set controlled gradients of NiCl2, a chemorepellent, in microchannels to quantify the motion of Escherichia coli over a broad range of concentrations. The experimental technique measured the motion of the bacteria in space and time and further related the motion to the local concentration profile of the repellent. Results show that the swimming speed of bacteria increases with an increasing concentration of repellent, which in turn enhances the drift velocity. The contribution of the increased swimming speed to the total drift velocity was in the range of 20 to 40%, with the remaining contribution coming from the modulation of the tumble frequency. A simple model that incorporates receptor dynamics, including adaptation, intracellular signaling, and swimming speed variation, was able to qualitatively capture the observed trend in drift velocity.

Escherichia coli modulates its motor speed on sensing an attractant.

It is well known that Escherichia coli achieves chemotaxis by modulating the bias of the flagellar motor. Recent experiments have shown that the bacteria vary their swimming speeds as well in presence of attractants. However, this increase in the swimming speed in response to the attractants has not been correlated with the increase in the flagellar motor speed. Using flickering dark-field microscopy, we measure the head-rotation speed of a large population of cells to correlate it with the flagellar motor speed. Experiments performed with wild-type and trg-deletion mutant strains suggest that the cells are capable of modulating the flagellar motor speed via mere sensing of a ligand. The motor speed can be further correlated with the swimming speed of the cells and was found to be linear. These results suggest the existence of a hitherto unknown intra-cellular pathway that modulates the flagellar motor speed in response to sensing of chemicals, thereby making chemotaxis more efficient than previously known.

Variation in swimming speed of Escherichia coli in response to attractant.

It is well known that Escherichia coli executes chemotactic motion in response to chemical cues by modulating the flagellar motor bias alone. However, previous studies have reported the possibility of variation in run speed in the presence of attractants although it is unclear whether bacteria can deliberately modulate their swimming speeds in response to environmental cues or if the motor speeds are hardwired. By studying the detailed motion of cells in a uniform concentration of glucose and its non-metabolizable analogue, we show that changing concentrations may be accompanied by variation in the swimming speed. For a fixed run duration, cells exposed to the attractants achieved a higher peak-swimming speed after a tumble compared with that in plain motility buffer. Our experiments using the mutant strain lacking the Trg sensor show no change in swimming speed with varying concentrations of the non-metabolizable analogue, suggesting that sensing may play a role in the observed variation of swimming speed.

A G-protein subunit translocation embedded network motif underlies GPCR regulation of calcium oscillations.

G-protein ßγ subunits translocate reversibly from the plasma membrane to internal membranes on receptor activation. Translocation rates differ depending on the γ subunit type. There is limited understanding of the role of the differential rates of Gßγ translocation in modulating signaling dynamics in a cell. Bifurcation analysis of the calcium oscillatory network structure predicts that the translocation rate of a signaling protein can regulate the damping of system oscillation. Here, we examined whether the Gßγ translocation rate regulates calcium oscillations induced by G-protein-coupled receptor activation. Oscillations in HeLa cells expressing γ subunit types with different translocation rates were imaged and quantitated. The results show that differential Gßγ translocation rates can underlie the diversity in damping characteristics of calcium oscillations among cells. Mathematical modeling shows that a translocation embedded motif regulates damping of G-protein-mediated calcium oscillations consistent with experimental data. The current study indicates that such a motif may act as a tuning mechanism to design oscillations with varying damping patterns by using intracellular translocation of a signaling component.

A systems biology-based mathematical model demonstrates the potential anti-stress effectiveness of a multi-nutrient botanical formulation.

Stress is an adaptive response to the stressors that adversely affects physiological and psychological health. Stress elicits HPA axis activation, resulting in cortisol release, ultimately contributing to oxidative, inflammatory, physiological and mental stress. Nutritional supplementations with antioxidant, anti-inflammatory, and stress-relieving properties are among widely preferred complementary approaches for the stress management. However, there is limited research on the potential combined impact of vitamins, minerals and natural ingredients on stress. In the present study, we have investigated the effect of a multi-nutrient botanical formulation, Nutrilite® Daily Plus, on clinical stress parameters. The stress-modulatory effects were quantified at population level using a customized sub-clinical inflammation mathematical model. The model suggested that combined intervention of botanical and micronutrients lead to significant decline in physical stress (75% decline), mental stress (70% decline), oxidative stress (55% decline) and inflammatory stress (75% decline) as evident from reduction in key stress parameters such as ROS, TNF-α, blood pressure, cortisol levels and PSS scores at both individual and population levels. Further, at the population level, the intervention relieved stress in 85% of individuals who moved towards a healthy state. The in silico studies strongly predicts the use of Gotukola based Nutrilite® Daily Plus as promising anti-stress formulation.

Characterization of burden on growth due to the nutritional state of media and pre-induced gene expression.

Studies have shown that the production of unnecessary proteins burdens the cellular growth mainly due to allocation of cellular resources to unnecessary protein synthesis, thereby limiting the resources available for growth. In the current study, we focus on the effect of pre-induction and nutritional status of the medium on the burden imposed on growth due to the synthesis of unnecessary protein. Escherichia coli cells with different history were grown in a glycerol media with and without IPTG to characterize the burden imposed due to the synthesis of ß-galactosidase. Effect of pre-induced lac operon on growth and ß-galactosidase expression on lactose milieu was also investigated. The study demonstrates that pre-induction has a strong influence on the extent of burden and is sustained in several generations. Further, the burden was much lower in a rich media relative to that observed in a minimal media.

Prediction by promoter logic in bacterial quorum sensing.

Quorum-sensing systems mediate chemical communication between bacterial cells, coordinating cell-density-dependent processes like biofilm formation and virulence-factor expression. In the proteobacterial LuxI/LuxR quorum sensing paradigm, a signaling molecule generated by an enzyme (LuxI) diffuses between cells and allosterically stimulates a transcriptional regulator (LuxR) to activate its cognate promoter (pR). By expressing either LuxI or LuxR in positive feedback from pR, these versatile systems can generate smooth (monostable) or abrupt (bistable) density-dependent responses to suit the ecological context. Here we combine theory and experiment to demonstrate that the promoter logic of pR - its measured activity as a function of LuxI and LuxR levels - contains all the biochemical information required to quantitatively predict the responses of such feedback loops. The interplay of promoter logic with feedback topology underlies the versatility of the LuxI/LuxR paradigm: LuxR and LuxI positive-feedback systems show dramatically different responses, while a dual positive/negative-feedback system displays synchronized oscillations. These results highlight the dual utility of promoter logic: to probe microscopic parameters and predict macroscopic phenotype.

A Systems Biology Approach To Disentangle the Direct and Indirect Effects of Global Transcription Factors on Gene Expression in Escherichia coli.

Delineating the pleiotropic effects of global transcriptional factors (TFs) is critical for understanding the system-wide regulatory response in a particular environment. Currently, with the availability of genome-wide TF binding and gene expression data for Escherichia coli, several gene targets can be assigned to the global TFs, albeit inconsistently. Here, using a systematic integrated approach with emphasis on metabolism, we characterized and quantified the direct effects as well as the growth rate-mediated indirect effects of global TFs using deletion mutants of FNR, ArcA, and IHF regulators (focal TFs) under glucose fermentative conditions. This categorization enabled us to disentangle the dense connections seen within the transcriptional regulatory network (TRN) and determine the exact nature of focal TF-driven epistatic interactions with other global and pathway-specific local regulators (iTFs). We extended our analysis to combinatorial deletions of these focal TFs to determine their cross talk effects as well as conserved patterns of regulatory interactions. Moreover, we predicted with high confidence several novel metabolite-iTF interactions using inferred iTF activity changes arising from the allosteric effects of the intracellular metabolites perturbed as a result of the absence of focal TFs. Further, using compendium level computational analyses, we revealed not only the coexpressed genes regulated by these focal TFs but also the coordination of the direct and indirect target expression in the context of the economy of intracellular metabolites. Overall, this study leverages the fundamentals of TF-driven regulation, which could serve as a better template for deciphering mechanisms underlying complex phenotypes. IMPORTANCE Understanding the pleiotropic effects of global TFs on gene expression and their relevance underlying a specific response in a particular environment has been challenging. Here, we distinguish the TF-driven direct effects and growth rate-mediated indirect effects on gene expression using single- and double-deletion mutants of FNR, ArcA, and IHF regulators under anaerobic glucose fermentation. Such dissection assists us in unraveling the precise nature of interactions existing between the focal TF(s) and several other TFs, including those altered by allosteric effects of intracellular metabolites. We were able to recapitulate the previously known metabolite-TF interactions and predict novel interactions with high confidence. Furthermore, we determined that the direct and indirect gene expression have a strong connection with each other when analyzed using the coexpressed- or coregulated-gene approach. Deciphering such regulatory patterns explicitly from the metabolism point of view would be valuable in understanding other unpredicted complex regulation existing in nature.

Effect of substrate and IPTG concentrations on the burden to growth of Escherichia coli on glycerol due to the expression of Lac proteins.

Expression of proteins unneeded for growth diverts cellular resources from making necessary protein and leads to a reduction in the growth rate of an organism. This reduction in growth rate is termed as cost. Cost plays an important role in determining the selected expression of a protein in a particular environment. Characterization of cost is important in biotechnology industries where microorganisms are used to produce foreign proteins. We have used the lactose system in Escherichia coli to quantify the cost of growth on glycerol in the presence of isopropyl-ß-D-thiogalactopyranoside (IPTG), an inducer of the lactose system. The effect of the concentration of the carbon source, glycerol, and the inducer of Lac enzymes, IPTG, is studied. The results show that the cost is dependent on the glycerol concentration with a decreasing trend with increasing concentration of glycerol. Also as expected, the cost increases and saturates at a higher concentration of IPTG. The studies also demonstrate that the cost is higher in early exponential phase relative to late exponential phase during the growth as has been reported in the literature. Hill equation fit yielded a typical Monod-type expression for growth on glycerol with and without IPTG. An apparent half-saturation constant was defined which was used to characterize the burden on growth due to protein expression.

Quantification of metabolism in Saccharomyces cerevisiae under hyperosmotic conditions using elementary mode analysis.

Yeast metabolism under hyperosmotic stress conditions was quantified using elementary mode analysis to obtain insights into the metabolic status of the cell. The fluxes of elementary modes were determined as solutions to a linear program that used the stoichiometry of the elementary modes as constraints. The analysis demonstrated that distinctly different sets of elementary modes operate under normal and hyperosmotic conditions. During the adaptation phase, elementary modes that only produce glycerol are active, while elementary modes that yield biomass, ethanol, and glycerol become active after the adaptive phase. The flux distribution in the metabolic network, calculated using the fluxes in the elementary modes, was employed to obtain the flux ratio at key nodes. At the glucose 6-phosphate (G6P) node, 25% of the carbon influx was diverted towards the pentose phosphate pathway under normal growth conditions, while only 0.3% of the carbon flux was diverted towards the pentose phosphate pathway during growth at 1 M NaCl, indicating that cell growth is arrested under hyperosmotic conditions. Further, objective functions were used in the linear program to obtain optimal solution spaces corresponding to the different accumulation rates. The analysis demonstrated that while biomass formation was optimal under normal growth conditions, glycerol synthesis was closer to optimal during adaptation to osmotic shock.

Global pleiotropic effects in adaptively evolved Escherichia coli lacking CRP reveal molecular mechanisms that define the growth physiology.

Evolution facilitates emergence of fitter phenotypes by efficient allocation of cellular resources in conjunction with beneficial mutations. However, system-wide pleiotropic effects that redress the perturbations to the apex node of the transcriptional regulatory networks remain unclear. Here, we elucidate that absence of global transcriptional regulator CRP in Escherichia coli results in alterations in key metabolic pathways under glucose respiratory conditions, favouring stress- or hedging-related functions over growth-enhancing functions. Further, we disentangle the growth-mediated effects from the CRP regulation-specific effects on these metabolic pathways. We quantitatively illustrate that the loss of CRP perturbs proteome efficiency, as evident from metabolic as well as ribosomal proteome fractions, that corroborated with intracellular metabolite profiles. To address how E. coli copes with such systemic defect, we evolved Δcrp mutant in the presence of glucose. Besides acquiring mutations in the promoter of glucose transporter ptsG, the evolved populations recovered the metabolic pathways to their pre-perturbed state coupled with metabolite re-adjustments, which altogether enabled increased growth. By contrast to Δcrp mutant, the evolved strains remodelled their proteome efficiency towards biomass synthesis, albeit at the expense of carbon efficiency. Overall, we comprehensively illustrate the genetic and metabolic basis of pleiotropic effects, fundamental for understanding the growth physiology.

Phenotypic characterization of Corynebacterium glutamicum under osmotic stress conditions using elementary mode analysis.

Corynebacterium glutamicum, a soil bacterium, is used to produce amino acids such as lysine and glutamate. C. glutamicum is often exposed to osmolality changes in its medium, and the bacterium has therefore evolved several adaptive response mechanisms to overcome them. In this study we quantify the metabolic response of C. glutamicum under osmotic stress using elementary mode analysis (EMA). Further, we obtain the optimal phenotypic space for the synthesis of lysine and formation of biomass. The analysis demonstrated that with increasing osmotic stress, the flux towards trehalose formation and energy-generating pathways increased, while the flux of anabolic reactions diminished. Nodal analysis indicated that glucose-6-phosphate, phosphoenol pyruvate, and pyruvate nodes were capable of adapting to osmotic stress, whereas the oxaloacetic acid node was relatively unresponsive. Fewer elementary modes were active under stress indicating the rigid behavior of the metabolism in response to high osmolality. Optimal phenotypic space analysis revealed that under normal conditions the organism optimized growth during the initial log phase and lysine and trehalose formation during the stationary phase. However, under osmotic stress, the analysis demonstrated that the organism operates under suboptimal conditions for growth, and lysine and trehalose formation.

Ligand sensing enhances bacterial flagellar motor output via stator recruitment.

It is well known that flagellated bacteria, such as Escherichia coli, sense chemicals in their environment by a chemoreceptor and relay the signals via a well-characterized signaling pathway to the flagellar motor. It is widely accepted that the signals change the rotation bias of the motor without influencing the motor speed. Here, we present results to the contrary and show that the bacteria is also capable of modulating motor speed on merely sensing a ligand. Step changes in concentration of non-metabolizable ligand cause temporary recruitment of stator units leading to a momentary increase in motor speeds. For metabolizable ligand, the combined effect of sensing and metabolism leads to higher motor speeds for longer durations. Experiments performed with mutant strains delineate the role of metabolism and sensing in the modulation of motor speed and show how speed changes along with changes in bias can significantly enhance response to changes in its environment.

Global Transcriptional Regulators Fine-Tune the Translational and Metabolic Efficiency for Optimal Growth of Escherichia coli.

Global transcriptional regulators coordinate complex genetic interactions that bestow better adaptability for an organism against external and internal perturbations. These transcriptional regulators are known to control an enormous array of genes with diverse functionalities. However, regulator-driven molecular mechanisms that underpin precisely tuned translational and metabolic processes conducive for rapid exponential growth remain obscure. Here, we comprehensively reveal the fundamental role of global transcriptional regulators FNR, ArcA, and IHF in sustaining translational and metabolic efficiency under glucose fermentative conditions in Escherichia coli By integrating high-throughput gene expression profiles and absolute intracellular metabolite concentrations, we illustrate that these regulators are crucial in maintaining nitrogen homeostasis, govern expression of otherwise unnecessary or hedging genes, and exert tight control on metabolic bottleneck steps. Furthermore, we characterize changes in expression and activity profiles of other coregulators associated with these dysregulated metabolic pathways, determining the regulatory interactions within the transcriptional regulatory network. Such systematic findings emphasize their importance in optimizing the proteome allocation toward metabolic enzymes as well as ribosomes, facilitating condition-specific phenotypic outcomes. Consequentially, we reveal that disruption of this inherent trade-off between ribosome and metabolic proteome economy due to the loss of regulators resulted in lowered growth rates. Moreover, our findings reinforce that the accumulations of intracellular metabolites in the event of proteome repartitions negatively affects the glucose uptake. Overall, by extending the three-partition proteome allocation theory concordant with multi-omics measurements, we elucidate the physiological consequences of loss of global regulators on central carbon metabolism restraining the organism to attain maximal biomass synthesis.IMPORTANCE Cellular proteome allocation in response to environmental or internal perturbations governs their eventual phenotypic outcome. This entails striking an effective balance between amino acid biosynthesis by metabolic proteins and its consumption by ribosomes. However, the global transcriptional regulator-driven molecular mechanisms that underpin their coordination remains unexplored. Here, we emphasize that global transcriptional regulators, known to control expression of a myriad of genes, are fundamental for precisely tuning the translational and metabolic efficiencies that define the growth optimality. Towards this, we systematically characterized the single deletion effect of FNR, ArcA, and IHF regulators of Escherichia coli on exponential growth under anaerobic glucose fermentative conditions. Their absence disrupts the stringency of proteome allocation, which manifests as impairment in key metabolic processes and an accumulation of intracellular metabolites. Furthermore, by incorporating an extension to the empirical growth laws, we quantitatively demonstrate the general design principles underlying the existence of these regulators in E. coli.