Juan Nogales - Academia.edu (original) (raw)

Papers by Juan Nogales

Microbial Cell Factories, 2013

Background: The metabolic capabilities of acetogens to ferment a wide range of sugars, to grow au... more Background: The metabolic capabilities of acetogens to ferment a wide range of sugars, to grow autotrophically on H 2 /CO 2 , and more importantly on synthesis gas (H 2 /CO/CO 2 ) make them very attractive candidates as production hosts for biofuels and biocommodities. Acetogenic metabolism is considered one of the earliest modes of bacterial metabolism. A thorough understanding of various factors governing the metabolism, in particular energy conservation mechanisms, is critical for metabolic engineering of acetogens for targeted production of desired chemicals. Results: Here, we present the genome-scale metabolic network of Clostridium ljungdahlii, the first such model for an acetogen. This genome-scale model (iHN637) consisting of 637 genes, 785 reactions, and 698 metabolites captures all the major central metabolic and biosynthetic pathways, in particular pathways involved in carbon fixation and energy conservation. A combination of metabolic modeling, with physiological and transcriptomic data provided insights into autotrophic metabolism as well as aided the characterization of a nitrate reduction pathway in C. ljungdahlii. Analysis of the iHN637 metabolic model revealed that flavin based electron bifurcation played a key role in energy conservation during autotrophic growth and helped identify genes for some of the critical steps in this mechanism. Conclusions: iHN637 represents a predictive model that recapitulates experimental data, and provides valuable insights into the metabolic response of C. ljungdahlii to genetic perturbations under various growth conditions. Thus, the model will be instrumental in guiding metabolic engineering of C. ljungdahlii for the industrial production of biocommodities and biofuels.

Applied and Environmental Microbiology, 2015

Thermotoga maritima is a hyperthermophilic anaerobe that utilizes a vast network of ABC transport... more Thermotoga maritima is a hyperthermophilic anaerobe that utilizes a vast network of ABC transporters to efficiently metabolize a variety of carbon sources to produce hydrogen. For unknown reasons, this organism does not metabolize glucose as readily as it does glucose di- and polysaccharides. The leading hypothesis implicates the thermolability of glucose at the physiological temperatures at which T. maritima lives. After a 25-day laboratory evolution, phenotypes were observed with growth rates up to 1.4 times higher than and glucose utilization rates exceeding 50% those of the wild type. Genome resequencing revealed mutations in evolved cultures related to glucose-responsive ABC transporters. The native glucose ABC transporter, GluEFK, has more abundant transcripts either as a result of gene duplication-amplification or through mutations to the operator sequence regulating this operon. Conversely, BglEFGKL, a transporter of beta-glucosides, is substantially downregulated due to a nonsense mutation to the solute binding protein or due to a deletion of the upstream promoter. Analysis of the ABC2 uptake porter families for carbohydrate and peptide transport revealed that the solute binding protein, often among the transcripts detected at the highest levels, is predominantly downregulated in the evolved cultures, while the membrane-spanning domain and nucleotide binding components are less varied. Similar trends were observed in evolved strains grown on glycerol, a substrate that is not dependent on ABC transporters. Therefore, improved growth on glucose is achieved through mutations favoring GluEFK expression over BglEFGKL, and in lieu of carbon catabolite repression, the ABC transporter network is modulated to achieve improved growth fitness.

Springer Protocols Handbooks, 2015

Springer Protocols Handbooks, 2014

ABSTRACT

2013 American Control Conference, 2013

An analytical framework for the study of a generic distribution problem is introduced in which a ... more An analytical framework for the study of a generic distribution problem is introduced in which a group of agents with different capabilities intend to maximize total utility by dividing themselves into various subgroups without any form of global information-sharing or centralized decision-making. The marginal utility of belonging to a particular subgroup rests on the well-known concept in economic theory of the law of diminishing returns. For a class of discrete event systems, we identify a set of conditions that define local information and cooperation requirements, and prove that if the proposed conditions are satisfied a stable agent distribution representing a Pareto optimum is achieved even under random but bounded decision and transition delays.

Current Opinion in Biotechnology, 2013

Our view on the bacterial responses to the aerobic degradation of aromatic compounds has been enr... more Our view on the bacterial responses to the aerobic degradation of aromatic compounds has been enriched considerably by the current omic methodologies and systems biology approaches, revealing the participation of intricate metabolic and regulatory networks. New enzymes, transporters, and specific/global regulatory systems have been recently characterized, and reveal that the widespread biodegradation capabilities extend to unexpected substrates such as lignin. A completely different biochemical strategy based on the formation of aryl-CoA epoxide intermediates has been unraveled for aerobic hybrid pathways, such as those involved in benzoate and phenylacetate degradation. Aromatic degradation pathways are also an important source of metabolic exchange factors and, therefore, they play a previously unrecognized biological role in cell-to-cell communication. Beyond the native bacterial biodegradation capabilities, pathway evolution as well as computational and synthetic biology approaches are emerging as powerful tools to design novel strain-specific pathways for degradation of xenobiotic compounds.

Molecular microbiology, 2011

Gallic acid (3,4,5-trihydroxybenzoic acid, GA) is widely distributed in nature, being a major phe... more Gallic acid (3,4,5-trihydroxybenzoic acid, GA) is widely distributed in nature, being a major phenolic pollutant and a commonly used antioxidant and building-block for drug development. We have characterized the first complete cluster (gal genes) responsible for growth in GA in a derivative of the model bacterium Pseudomonas putida KT2440. GalT mediates specific GA uptake and chemotaxis, and highlights the critical role of GA transport in bacterial adaptation to GA consumption. The proposed GA degradation via the central intermediate 4-oxalomesaconic acid (OMA) was revisited and all enzymes involved have been identified. Thus, GalD is the prototype of a new subfamily of isomerases that catalyses a biochemical step that remained unknown, i.e. the tautomerization of the OMAketo generated by the GalA dioxygenase to OMAenol. GalB is the founding member of a new family of zinc-containing hydratases that converts OMAenol into 4-carboxy-4-hydroxy-2-oxoadipic acid (CHA). galC encodes the al...

Genome announcements, 2015

We report here the draft genome sequence of Pseudomonas azelaica strain Aramco J (7.3 Mbp; GC con... more We report here the draft genome sequence of Pseudomonas azelaica strain Aramco J (7.3 Mbp; GC content, 61.9%), one of the few bacteria that can completely mineralize different hydroxybiphenyls, e.g., 2-hydroxybiphenyl, 2,2'-dihydroxybiphenyl, and 3-hydroxybiphenyl. The findings obtained from its genome annotation suggest that this strain becomes a useful biocatalyst for aromatic bioconversions.

BMC Systems Biology, 2008

Background: Pseudomonas putida is the best studied pollutant degradative bacteria and is harnesse... more Background: Pseudomonas putida is the best studied pollutant degradative bacteria and is harnessed by industrial biotechnology to synthesize fine chemicals. Since the publication of P. putida KT2440's genome, some in silico analyses of its metabolic and biotechnology capacities have been published. However, global understanding of the capabilities of P. putida KT2440 requires the construction of a metabolic model that enables the integration of classical experimental data along with genomic and high-throughput data. The constraint-based reconstruction and analysis (COBRA) approach has been successfully used to build and analyze in silico genome-scale metabolic reconstructions.

Journal of Biological Chemistry, 2005

Nature Biotechnology, 2014

Proceedings of the National Academy of Sciences, 2012

Photosynthesis has recently gained considerable attention for its potential role in the developme... more Photosynthesis has recently gained considerable attention for its potential role in the development of renewable energy sources. Optimizing photosynthetic organisms for biomass or biofuel production will therefore require a systems understanding of photosynthetic processes. We reconstructed a high-quality genome-scale metabolic network for Synechocystis sp. PCC6803 that describes key photosynthetic processes in mechanistic detail. We performed an exhaustive in silico analysis of the reconstructed photosynthetic process under different light and inorganic carbon (Ci) conditions as well as under genetic perturbations. Our key results include the following. (i) We identified two main states of the photosynthetic apparatus: a Ci-limited state and a light-limited state. (ii) We discovered nine alternative electron flow pathways that assist the photosynthetic linear electron flow in optimizing the photosynthesis performance. (iii) A high degree of cooperativity between alternative pathways was found to be critical for optimal autotrophic metabolism. Although pathways with high photosynthetic yield exist for optimizing growth under suboptimal light conditions, pathways with low photosynthetic yield guarantee optimal growth under excessive light or Ci limitation. (iv) Photorespiration was found to be essential for the optimal photosynthetic process, clarifying its role in high-light acclimation. Finally, (v) an extremely high photosynthetic robustness drives the optimal autotrophic metabolism at the expense of metabolic versatility and robustness. The results and modeling approach presented here may promote a better understanding of the photosynthetic process. They can also guide bioengineering projects toward optimal biofuel production in photosynthetic organisms.

Genome Announcements, 2014

Pseudomonas azelaica HBP1 (DSM 8897) is one of the few bacteria able to completely mineralize the... more Pseudomonas azelaica HBP1 (DSM 8897) is one of the few bacteria able to completely mineralize the 2-hydroxybiphenyl biocide. Here, we report the draft genome sequence of this strain (7.4 Mbp; G؉C content, 63.5%) and the findings obtained from its genome annotation.

Systematic and Applied Microbiology, 2015

The genomic features of Azoarcus sp. CIB reflect its most distinguishing phenotypes as a diazotro... more The genomic features of Azoarcus sp. CIB reflect its most distinguishing phenotypes as a diazotroph, facultative anaerobe, capable of degrading either aerobically and/or anaerobically a wide range of aromatic compounds, including some toxic hydrocarbons such as toluene and m-xylene, as well as its endophytic lifestyle. The analyses of its genome have expanded the catabolic potential of strain CIB toward common natural compounds, such as certain diterpenes, that were not anticipated as carbon sources. The high number of predicted solvent efflux pumps and heavy metal resistance gene clusters has provided the first evidence for two environmentally relevant features of this bacterium that remained unknown. Genome mining has revealed several gene clusters likely involved in the endophytic lifestyle of strain CIB, opening the door to the molecular characterization of some plant growth promoting traits. Horizontal gene transfer and mobile genetic elements appear to have played a major role as a mechanism of adaptation of this bacterium to different lifestyles. This work paves the way for a systems biology-based understanding of the abilities of Azoarcus sp. CIB to integrate aerobic and anaerobic metabolism of aromatic compounds, tolerate stress conditions, and interact with plants as an endophyte of great potential for phytostimulation and phytoremediation strategies. Comparative genomics provides an Azoarcus pan genome that confirms the global metabolic flexibility of this genus, and suggests that its phylogeny should be revisited.

Microbiology, 2007

Phenylacetic acid (PA) degradation in bacteria involves an aerobic hybrid pathway encoded by the ... more Phenylacetic acid (PA) degradation in bacteria involves an aerobic hybrid pathway encoded by the paa gene cluster. It is shown here that succinyl-CoA is one of the final products of this pathway in Pseudomonas putida and Escherichia coli. Moreover, in vivo and in vitro studies revealed that the paaE gene encodes the b-ketoadipyl-CoA thiolase that catalyses the last step of the PA catabolic pathway, i.e. the thiolytic cleavage of b-ketoadipyl-CoA to succinyl-CoA and acetyl-CoA. Succinyl-CoA is suggested as a common final product of aerobic hybrid pathways devoted to the catabolism of aromatic compounds.

Environmental Microbiology, 2006

Bacterial transcriptional networks are built on a hierarchy of regulators, on top of which lie th... more Bacterial transcriptional networks are built on a hierarchy of regulators, on top of which lie the components of the RNA polymerase (in particular the sigma factors) and the global control elements, which play a pivotal role. We have designed a genome-wide oligonucleotide-based DNA microarray for Pseudomonas putida KT2440. In combination with real-time reverse transcription polymerase chain reaction (RT-PCR), we have used it to analyse the expression pattern of the genes encoding the RNA polymerase subunits (the core enzyme and the 24 sigma factors), and various proteins involved in global regulation (Crc, Lrp, Fur, Anr, Fis, CsrA, IHF, HupA, HupB, HupN, BipA and several MvaT-like proteins), during the shift from exponential growth in rich medium into starvation and stress brought about by the entry into stationary phase. Expression of the genes encoding the RNA polymerase core and the vegetative sigma factor decreased in stationary phase, while that of s S increased. Data obtained for s N , s H , FliA and for the 19 extracytoplasmic function (ECF)-like sigma factors suggested that their mRNA levels change little upon entry into stationary phase. Expression of Crc, BipA, Fis, HupB, HupN and the MvaT-like protein PP3693 decreased in stationary phase, while that of HupA and the MvaT-like protein PP3765 increased significantly. Expression of IHF was indicative of posttranscriptional control. These results provide the first global study of the expression of the transcriptional machinery through the exponential stationary-phase shift in P. putida .

Environmental microbiology, Jan 25, 2015

In metabolically versatile bacteria, carbon catabolite repression (CCR) facilitates the preferent... more In metabolically versatile bacteria, carbon catabolite repression (CCR) facilitates the preferential assimilation of the most efficient carbon sources, improving growth rates and fitness. In Pseudomonas putida, the Crc and Hfq proteins and the CrcZ and CrcY small RNAs, which are believed to antagonize Crc/Hfq, are key players in CCR. Unlike that seen in other bacterial species, succinate and glucose elicit weak CCR in this bacterium. In the present work, metabolic, transcriptomic and constraint-based metabolic flux analyses were combined to clarify whether P. putida prefers succinate or glucose, and to identify the role of the Crc protein in the metabolism of these compounds. When provided simultaneously, succinate was consumed faster than glucose, although both compounds were metabolized. CrcZ and CrcY levels were lower when both substrates were present than when only one was provided, suggesting a role for Crc in coordinating metabolism of these compounds. Flux distribution analys...

International Journal of Hydrogen Energy, 2012

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Mol. BioSyst., 2014

The increasing need to replace oil-based products and to address global climate change concerns h... more The increasing need to replace oil-based products and to address global climate change concerns has triggered considerable interest in photosynthetic microorganisms. Cyanobacteria, in particular, have great potential as biocatalysts for fuels and fine-chemicals. During the last few years the biotechnological applications of cyanobacteria have experienced an unprecedented increase and the use of these photosynthetic organisms for chemical production is becoming a tangible reality. However, the field is still immature and many concerns about the economic feasibility of the biotechnological potential of cyanobacteria remain. In this review we describe recent successes in biofuel and fine-chemical production using cyanobacteria. We discuss the role of the photosynthetic metabolism and highlight the need for systems-level metabolic optimization in order to achieve the true potential of cyanobacterial biocatalysts.

Bioengineered, 2013

We recently assessed the metabolism of Synechocystis sp PCC6803 through a constraints-based recon... more We recently assessed the metabolism of Synechocystis sp PCC6803 through a constraints-based reconstruction and analysis approach and identified its main metabolic properties. These include reduced metabolic robustness, in contrast to a high photosynthetic robustness driving the optimal autotrophic metabolism. Here, we address how these metabolic features affect biotechnological capabilities of this bacterium. The search for growth-coupled overproducer strains revealed that the carbon flux re-routing, but not the electron flux, is significantly more challenging under autotrophic conditions than under mixo- or heterotrophic conditions. We also found that the blocking of the light-driven metabolism was required for carbon flux re-routing under mixotrophic conditions. Overall, our analysis, which represents the first systematic evaluation of the biotechnological capabilities of a photosynthetic organism, paradoxically suggests that the light-driven metabolism itself and its unique metabolic features are the main bottlenecks in harnessing the biotechnological potential of Synechocystis.