scholarly journals Massively Parallel Fitness Profiling Reveals Multiple Novel Enzymes in Pseudomonas putida Lysine Metabolism

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
Mitchell G. Thompson ◽  
Jacquelyn M. Blake-Hedges ◽  
Pablo Cruz-Morales ◽  
Jesus F. Barajas ◽  
Samuel C. Curran ◽  
...  
Keyword(s):  
Metabolic Pathway ◽  
Massively Parallel ◽  
Central Metabolism ◽  
It Use ◽  
Content Type ◽  
Biochemical Function ◽  
Commodity Chemicals ◽  
Domains Of Life ◽  
Lysine Catabolism ◽  
Lysine Metabolism

P. putidalysine metabolism can produce multiple commodity chemicals, conferring great biotechnological value. Despite much research, the connection of lysine catabolism to central metabolism inP. putidaremained undefined. Here, we used random barcode transposon sequencing to fill the gaps of lysine metabolism inP. putida. We describe a route of 2-oxoadipate (2OA) catabolism, which utilizes DUF1338-containing proteinP. putida5260 (PP_5260) in bacteria. Despite its prevalence in many domains of life, DUF1338-containing proteins have had no known biochemical function. We demonstrate that PP_5260 is a metalloenzyme which catalyzes an unusual route of decarboxylation of 2OA tod-2-hydroxyglutarate (d-2HG). Our screen also identified a recently described novel glutarate metabolic pathway. We validate previous results and expand the understanding of glutarate hydroxylase CsiD by showing that can it use either 2OA or 2KG as a cosubstrate. Our work demonstrated that biological novelty can be rapidly identified using unbiased experimental genetics and that RB-TnSeq can be used to rapidly validate previous results.

scholarly journals Massively parallel fitness profiling reveals multiple novel enzymes inPseudomonas putidalysine metabolism

2018 ◽  
Author(s):  
Mitchell G. Thompson ◽  
Jacquelyn M. Blake-Hedges ◽  
Pablo Cruz-Morales ◽  
Jesus F. Barajas ◽  
Samuel C. Curran ◽  
...  
Keyword(s):  
Tca Cycle ◽  
Central Metabolism ◽  
It Use ◽  
Catabolic Genes ◽  
Genome Wide ◽  
The Tca Cycle ◽  
A Genome ◽  
Domains Of Life ◽  
Lysine Degradation ◽  
Lysine Metabolism

AbstractDespite intensive study for 50 years, the biochemical and genetic links between lysine metabolism and central metabolism inPseudomonas putidaremain unresolved. To establish these biochemical links, we leveraged Random Barcode Transposon Sequencing (RB-TnSeq), a genome-wide assay measuring the fitness of thousands of genes in parallel, to identify multiple novel enzymes in both L- and D-lysine metabolism. We first describe three pathway enzymes that catabolize L-2-aminoadipate (L-2AA) to 2-ketoglutarate (2KG), connecting D-lysine to the TCA cycle. One of these enzymes, PP_5260, contains a DUF1338 domain, a family with no previously described biological function. Our work also identified the recently described CoA independent route of L-lysine degradation that metabolizes to succinate. We expanded on previous findings by demonstrating that glutarate hydroxylase CsiD is promiscuous in its 2-oxoacid selectivity. Proteomics of select pathway enzymes revealed that expression of catabolic genes is highly sensitive to particular pathway metabolites, implying intensive local and global regulation. This work demonstrates the utility of RB-TnSeq for discovering novel metabolic pathways in even well-studied bacteria, as well as a powerful tool for validating previous research.ImportanceP. putidalysine metabolism can produce multiple commodity chemicals, conferring great biotechnological value. Despite much research, connecting lysine catabolism to central metabolism inP. putidaremained undefined. Herein we use Random Barcode Transposon Sequencing to fill in the gaps of lysine metabolism inP. putida. We describe a route of 2-oxoadipate (2OA) catabolism in bacteria, which utilizes DUF1338 containing protein PP_5260. Despite its prevalence in many domains of life, DUF1338 containing proteins had no known biochemical function. We demonstrate PP_5260 is a metalloenzyme which catalyzes an unusual 2OA to D-2HG decarboxylation. Our screen also identified a recently described novel glutarate metabolic pathway. We validate previous results, and expand the understanding of glutarate hydroxylase CsiD by showing can it use either 2OA or 2KG as a cosubstrate. Our work demonstrates biological novelty can be rapidly identified using unbiased experimental genetics, and that RB-TnSeq can be used to rapidly validate previous results.

Configurational examination of successful human resources outsourcing

2017 ◽  
Vol 55 (8) ◽  
pp. 1681-1697
Author(s):  
Thuy Thi Thanh Nguyen ◽  
Man-Ling Chang
Keyword(s):  
Human Resources ◽  
Design Methodology ◽  
Qualitative Comparative Analysis ◽  
Task Interdependence ◽  
It Use ◽  
Research Attention ◽  
Content Type ◽  
Provider Quality ◽  
Core Condition ◽  
Practical Implications

Purpose The causal conditions leading to successful human resources outsourcing (HRO) have until now attracted only limited research attention. To address this shortcoming, the purpose of this paper is to investigate how a firm and a HR provider should work together to ensure successful HRO. Design/methodology/approach By applying a fuzzy-set qualitative comparative analysis, the authors demonstrate how combining in different ways provider quality, business understanding, trust, and closed HRO networks can lead to HRO success. In addition, this study compares the causal combinations between cases with differing levels of HR-task interdependence and IT use. Findings The results of this study reveal three common solutions that lead to successful HRO from both economic and relational viewpoints. The findings also indicate that, from an economic viewpoint, trust is a core condition for achieving HRO success. Moreover, HR-task interdependence and IT use alter the number of solutions and the conditions for attaining HRO success. Practical implications Based on the solutions proposed herein, firms can consider different ways to achieve HRO success under diverse conditions. Originality/value This work also contributes to building new theories regarding HRO, trust, knowledge sharing, and IT use.

scholarly journals Arabinose-Induced Catabolite Repression as a Mechanism for Pentose Hierarchy Control inClostridium acetobutylicumATCC 824

mSystems ◽  
2018 ◽  
Vol 3 (5) ◽  
Author(s):  
Matthew D. Servinsky ◽  
Rebecca L. Renberg ◽  
Matthew A. Perisin ◽  
Elliot S. Gerlach ◽  
Sanchao Liu ◽  
...  
Keyword(s):  
Transcriptional Regulation ◽  
Genetic Engineering ◽  
Catabolite Repression ◽  
Clostridium Acetobutylicum ◽  
Regulation Mechanism ◽  
Mrna Levels ◽  
Chemical Production ◽  
Content Type ◽  
Commodity Chemicals ◽  
Pentose Utilization

ABSTRACTBacterial fermentation of carbohydrates from sustainable lignocellulosic biomass into commodity chemicals by the anaerobic bacteriumClostridium acetobutylicumis a promising alternative source to fossil fuel-derived chemicals. Recently, it was demonstrated that xylose is not appreciably fermented in the presence of arabinose, revealing a hierarchy of pentose utilization in this organism (L. Aristilde, I. A. Lewis, J. O. Park, and J. D. Rabinowitz, Appl Environ Microbiol 81:1452–1462, 2015,https://doi.org/10.1128/AEM.03199-14). The goal of the current study is to characterize the transcriptional regulation that occurs and perhaps drives this pentose hierarchy. Carbohydrate consumption rates showed that arabinose, like glucose, actively represses xylose utilization in cultures fermenting xylose. Further, arabinose addition to xylose cultures led to increased acetate-to-butyrate ratios, which indicated a transition of pentose catabolism from the pentose phosphate pathway to the phosphoketolase pathway. Transcriptome sequencing (RNA-Seq) confirmed that arabinose addition to cells actively growing on xylose resulted in increased phosphoketolase (CA_C1343) mRNA levels, providing additional evidence that arabinose induces this metabolic switch. A significant overlap in differentially regulated genes after addition of arabinose or glucose suggested a common regulation mechanism. A putative open reading frame (ORF) encoding a potential catabolite repression phosphocarrier histidine protein (Crh) was identified that likely participates in the observed transcriptional regulation. These results substantiate the claim that arabinose is utilized preferentially over xylose inC. acetobutylicumand suggest that arabinose can activate carbon catabolite repression via Crh. Furthermore, they provide valuable insights into potential mechanisms for altering pentose utilization to modulate fermentation products for chemical production.IMPORTANCEClostridium acetobutylicumcan ferment a wide variety of carbohydrates to the commodity chemicals acetone, butanol, and ethanol. Recent advances in genetic engineering have expanded the chemical production repertoire ofC. acetobutylicumusing synthetic biology. Due to its natural properties and genetic engineering potential, this organism is a promising candidate for converting biomass-derived feedstocks containing carbohydrate mixtures to commodity chemicals via natural or engineered pathways. Understanding how this organism regulates its metabolism during growth on carbohydrate mixtures is imperative to enable control of synthetic gene circuits in order to optimize chemical production. The work presented here unveils a novel mechanism via transcriptional regulation by a predicted Crh that controls the hierarchy of carbohydrate utilization and is essential for guiding robust genetic engineering strategies for chemical production.

scholarly journals Physiology, Biochemistry, and Applications of F420- and Fo-Dependent Redox Reactions

2016 ◽  
Vol 80 (2) ◽  
pp. 451-493 ◽  
Author(s):  
Chris Greening ◽  
F. Hafna Ahmed ◽  
A. Elaaf Mohamed ◽  
Brendon M. Lee ◽  
Gunjan Pandey ◽  
...  
Keyword(s):  
Redox Reactions ◽  
Methanogenic Archaea ◽  
Content Type ◽  
Sulfate Reducing ◽  
Single Function ◽  
Wide Range ◽  
Domains Of Life ◽  
Photochemical Properties ◽  
Endogenous Metabolites ◽  
Dependent Processes

SUMMARY5-Deazaflavin cofactors enhance the metabolic flexibility of microorganisms by catalyzing a wide range of challenging enzymatic redox reactions. While structurally similar to riboflavin, 5-deazaflavins have distinctive and biologically useful electrochemical and photochemical properties as a result of the substitution of N-5 of the isoalloxazine ring for a carbon. 8-Hydroxy-5-deazaflavin (Fo) appears to be used for a single function: as a light-harvesting chromophore for DNA photolyases across the three domains of life. In contrast, its oligoglutamyl derivative F420is a taxonomically restricted but functionally versatile cofactor that facilitates many low-potential two-electron redox reactions. It serves as an essential catabolic cofactor in methanogenic, sulfate-reducing, and likely methanotrophic archaea. It also transforms a wide range of exogenous substrates and endogenous metabolites in aerobic actinobacteria, for example mycobacteria and streptomycetes. In this review, we discuss the physiological roles of F420in microorganisms and the biochemistry of the various oxidoreductases that mediate these roles. Particular focus is placed on the central roles of F420in methanogenic archaea in processes such as substrate oxidation, C1pathways, respiration, and oxygen detoxification. We also describe how two F420-dependent oxidoreductase superfamilies mediate many environmentally and medically important reactions in bacteria, including biosynthesis of tetracycline and pyrrolobenzodiazepine antibiotics by streptomycetes, activation of the prodrugs pretomanid and delamanid byMycobacterium tuberculosis, and degradation of environmental contaminants such as picrate, aflatoxin, and malachite green. The biosynthesis pathways of Foand F420are also detailed. We conclude by considering opportunities to exploit deazaflavin-dependent processes in tuberculosis treatment, methane mitigation, bioremediation, and industrial biocatalysis.

Emotional capability: the missing link between information technology capabilities and innovation performanceCapacidad emocional: el vínculo perdido entre capacidades de tecnologías de la información y desempeño innovadorCapacidade emocional: O elo perdido entre as capacidades das tecnologias da informação e o desempenho inovador

2021 ◽  
Vol ahead-of-print (ahead-of-print) ◽  
Author(s):  
José Arias-Pérez ◽  
Joaquin Alegre ◽  
Cristina Villar
Keyword(s):  
Information Technology ◽  
Information Processing ◽  
Statistical Significance ◽  
Organizational Factors ◽  
Structural Equations ◽  
Mediating Effect ◽  
It Services ◽  
It Use ◽  
Content Type ◽  
Strategic Market

Purpose There has been a great interest in the literature in understanding the incidence of information technology capabilities (ITC) on innovation performance (IP). Recently, it has been proven that this relationship is mediated by organizational factors requiring an additional effort in terms of information processing in a rational and analytical manner, including strategic market orientation and absorptive capacity; however, the role of emotions in this discussion has not been widely addressed. A scenario in which emotions are inhibited, in particular, emotional capability (EC), prevents the activation of other forms of cognition relating to intuition, experience and empathy. Therefore, the purpose of this paper is to analyze the mediating effect of EC on the relationship between ITC and IP. Design/methodology/approach The authors analyzed the statistical significance of the indirect effects through structural equations. Findings The results show the existence of partial mediation of EC. Therefore, it is evidenced that less rational constructs, such as EC, serve as a bridge between ITC and IP, on account of the improvement in information processing, the level of information technology (IT) use and virtual communication. Originality/value This finding is quite significant because it forces innovation researchers and practitioners to reconsider the prevailing study perspective that explores only analytical mediators, which implies an additional effort in terms of rational processing of information. All this because there is a risk of accentuating the cognitive overload and increasing the levels of stress that lead employees to stop receiving and using information provided by IT services, which compromises the possibility of obtaining better results in innovation.

scholarly journals Cofactor Selectivity in Methylmalonyl Coenzyme A Mutase, a Model Cobamide-Dependent Enzyme

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Olga M. Sokolovskaya ◽  
Kenny C. Mok ◽  
Jong Duk Park ◽  
Jennifer L. A. Tran ◽  
Kathryn A. Quanstrom ◽  
...  
Keyword(s):  
Binding Affinity ◽  
Bacterial Growth ◽  
Microbial Interactions ◽  
Vitamin B ◽  
Content Type ◽  
Domains Of Life ◽  
Enzyme Cofactors ◽  
Lower Ligand ◽  
Enzyme Selectivity

ABSTRACT Cobamides, a uniquely diverse family of enzyme cofactors related to vitamin B12, are produced exclusively by bacteria and archaea but used in all domains of life. While it is widely accepted that cobamide-dependent organisms require specific cobamides for their metabolism, the biochemical mechanisms that make cobamides functionally distinct are largely unknown. Here, we examine the effects of cobamide structural variation on a model cobamide-dependent enzyme, methylmalonyl coenzyme A (CoA) mutase (MCM). The in vitro binding affinity of MCM for cobamides can be dramatically influenced by small changes in the structure of the lower ligand of the cobamide, and binding selectivity differs between bacterial orthologs of MCM. In contrast, variations in the lower ligand have minor effects on MCM catalysis. Bacterial growth assays demonstrate that cobamide requirements of MCM in vitro largely correlate with in vivo cobamide dependence. This result underscores the importance of enzyme selectivity in the cobamide-dependent physiology of bacteria. IMPORTANCE Cobamides, including vitamin B12, are enzyme cofactors used by organisms in all domains of life. Cobamides are structurally diverse, and microbial growth and metabolism vary based on cobamide structure. Understanding cobamide preference in microorganisms is important given that cobamides are widely used and appear to mediate microbial interactions in host-associated and aquatic environments. Until now, the biochemical basis for cobamide preferences was largely unknown. In this study, we analyzed the effects of the structural diversity of cobamides on a model cobamide-dependent enzyme, methylmalonyl-CoA mutase (MCM). We found that very small changes in cobamide structure could dramatically affect the binding affinity of cobamides to MCM. Strikingly, cobamide-dependent growth of a model bacterium, Sinorhizobium meliloti, largely correlated with the cofactor binding selectivity of S. meliloti MCM, emphasizing the importance of cobamide-dependent enzyme selectivity in bacterial growth and cobamide-mediated microbial interactions.

Understanding the Chinese information culture and its impact on IT perceptions

2019 ◽  
Vol 33 (1) ◽  
pp. 56-82 ◽  
Author(s):  
Leida Chen ◽  
Xiaohan Zhang ◽  
Zhensheng Li
Keyword(s):  
Cultural Values ◽  
Information Science ◽  
Information Source ◽  
Information Control ◽  
Information Culture ◽  
It Use ◽  
Content Type ◽  
Individual Level ◽  
National Information ◽  
The Impact

Purpose Drawing upon the theory of information culture from information science and prior research on Chinese information practices. The purpose of this paper is to posit that instead of over-relying on Hofstede’s national cultural values, the IS field should develop theories on national information culture to bridge the theoretical gaps found in explaining the differences in individuals’ IT perceptions and behaviors across cultures. Design/methodology/approach To achieve the objectives of this study, two quantitative studies were conducted. The first study is exploratory in nature and focuses on the development and calibration of an instrument to measure the Chinese information culture (CIC). The second study takes a confirmatory approach to investigate the impact of the CIC on IT perceptions of manager. Findings The study develops and validates an instrument for measuring the CIC that comprises information source and format preference, information ownership and analysis style, and centralized decision-making and information control. The CIC is found to influence IT perceptions of managers including perceived performance improvement resulted from IT use, attitude toward using work technologies and attitude on the value of IT in general. Originality/value This study is one of the first studies to theorize national information culture to understand IT perceptions at the individual level. The development of the CIC measure provides the much needed methodological support for contextualizing cross-cultural research.

Exploring adaptive information sharing from the perspective of cognitive switching

2019 ◽  
Vol 71 (4) ◽  
pp. 535-557 ◽  
Author(s):  
Xianjin Zha ◽  
Haijuan Yang ◽  
Yalan Yan ◽  
Guanxiang Yan ◽  
Chengsong Huang ◽  
...  
Keyword(s):  
Information Sharing ◽  
Structural Equation ◽  
Information Technologies ◽  
Equation Modeling ◽  
Research Model ◽  
It Use ◽  
Content Type ◽  
Technology System ◽  
Use Behavior ◽  
Cognitive Switching

Purpose Microblogging as one kind of social media application provides an important information sharing platform. Adaptive information sharing is the combination of adaptive information technologies (IT) use behavior and information sharing behavior and subsequently refers to adaptive use of IT oriented to information sharing. The purpose of this paper is to understand adaptive information sharing in the context of microblogging from the perspective of cognitive switching. Design/methodology/approach A research model was developed and survey data were collected. The partial least squares structural equation modeling was employed to verify the research model. Findings Adaptive information sharing is positively impacted by other people’s use, discrepancies and deliberate initiatives among which other people’s use is the key determinant. Meanwhile, task self-efficacy positively moderates the effect of other people’s use on adaptive information sharing. Practical implications Developers of microblogging should as far as possible create learning atmosphere and learning culture. With learning atmosphere and culture, more and more users could keep on learning from observing other people. Consequently, more and more users would be willing to try new features of microblogging to share information. Originality/value This study examines adaptive information sharing by extending adaptive IT use behavior from the levels of technology, system and feature to the information level, presenting a new lens for adaptive IT use and information sharing alike.

scholarly journals Quantifying the Survival of MultipleSalmonella entericaSerovarsIn Vivovia Massively Parallel Whole-Genome Sequencing To Predict Zoonotic Risk

2017 ◽  
Vol 84 (4) ◽  
Author(s):  
Prerna Vohra ◽  
Marie Bugarel ◽  
Frances Turner ◽  
Guy H. Loneragan ◽  
Jayne C. Hope ◽  
...  
Keyword(s):  
Lymph Nodes ◽  
Genome Sequencing ◽  
Lymphatic System ◽  
Massively Parallel ◽  
Whole Genome ◽  
Content Type ◽  
Peripheral Lymph ◽  
Zoonotic Risk ◽  
Mixed Populations

ABSTRACTSalmonella entericais an animal and zoonotic pathogen of worldwide importance.Salmonellaserovars that differ in their host and tissue tropisms exist. Cattle are an important reservoir of human nontyphoidal salmonellosis, and contaminated bovine peripheral lymph nodes enter the food chain via ground beef. The relative abilities of different serovars to survive within the bovine lymphatic system are poorly understood and constrain the development of control strategies. This problem was addressed by developing a massively parallel whole-genome sequencing method to study mixed-serovar infectionsin vivo.Salmonellaserovars differ genetically by naturally occurring single nucleotide polymorphisms (SNPs) in certain genes. It was hypothesized that these SNPs could be used as markers to simultaneously identify serovars in mixed populations and quantify the abundance of each member in a population. The performance of the method was validatedin vitrousing simulated pools containing up to 11 serovars in various proportions. It was then applied to study serovar survivalin vivoin cattle challenged orally with the same 11 serovars. All the serovars successfully colonized the bovine lymphatic system, including the peripheral lymph nodes, and thus pose similar risks of zoonosis. This method enables the fates of multiple genetically unmodified strains to be evaluated simultaneously in a single animal. It could be useful in reducing the number of animals required to study mixed-strain infections and in testing the cross-protective efficacy of vaccines and treatments. It also has the potential to be applied to diverse bacterial species which possess shared but polymorphic alleles.IMPORTANCEWhile someSalmonellaserovars are more frequently isolated from lymph nodes rather than the feces and environment of cattle, the relative abilities of serovars to survive within the lymphatic system of cattle remain ill defined. A sequencing-based method which used available information from sequencedSalmonellagenomes to study the dynamics of mixed-serovar infectionsin vivowas developed. The main advantages of the method include the simultaneous identification and quantification of multiple strains without any genetic modification and minimal animal use. This approach could be used in vaccination trials or in epidemiological surveys where an understanding of the dynamics of closely related strains of a pathogen in mixed populations could inform the prediction of zoonotic risk and the development of intervention strategies.

scholarly journals Conversion of 4-Hydroxybutyrate to Acetyl Coenzyme A and Its Anapleurosis in the Metallosphaera sedula 3-Hydroxypropionate/4-Hydroxybutyrate Carbon Fixation Pathway

2014 ◽  
Vol 80 (8) ◽  
pp. 2536-2545 ◽  
Author(s):  
Aaron B. Hawkins ◽  
Michael W. W. Adams ◽  
Robert M. Kelly
Keyword(s):  
Carbon Fixation ◽  
Coenzyme A ◽  
Flux Analysis ◽  
Central Metabolism ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Metallosphaera Sedula ◽  
Acetyl Coenzyme ◽  
Coa Ligase

ABSTRACTThe extremely thermoacidophilic archaeonMetallosphaera sedula(optimum growth temperature, 73°C, pH 2.0) grows chemolithoautotrophically on metal sulfides or molecular hydrogen by employing the 3-hydroxypropionate/4-hydroxybutyrate (3HP/4HB) carbon fixation cycle. This cycle adds two CO2molecules to acetyl coenzyme A (acetyl-CoA) to generate 4HB, which is then rearranged and cleaved to form two acetyl-CoA molecules. Previous metabolic flux analysis showed that two-thirds of central carbon precursor molecules are derived from succinyl-CoA, which is oxidized to malate and oxaloacetate. The remaining one-third is apparently derived from acetyl-CoA. As such, the steps beyond succinyl-CoA are essential for completing the carbon fixation cycle and for anapleurosis of acetyl-CoA. Here, the final four enzymes of the 3HP/4HB cycle, 4-hydroxybutyrate-CoA ligase (AMP forming) (Msed_0406), 4-hydroxybutyryl-CoA dehydratase (Msed_1321), crotonyl-CoA hydratase/(S)-3-hydroxybutyryl-CoA dehydrogenase (Msed_0399), and acetoacetyl-CoA β-ketothiolase (Msed_0656), were produced recombinantly inEscherichia coli, combinedin vitro, and shown to convert 4HB to acetyl-CoA. Metabolic pathways connecting CO2fixation and central metabolism were examined using a gas-intensive bioreactor system in whichM. sedulawas grown under autotrophic (CO2-limited) and heterotrophic conditions. Transcriptomic analysis revealed the importance of the 3HP/4HB pathway in supplying acetyl-CoA to anabolic pathways generating intermediates inM. sedulametabolism. The results indicated that flux between the succinate and acetyl-CoA branches in the 3HP/4HB pathway is governed by 4-hydroxybutyrate-CoA ligase, possibly regulated posttranslationally by the protein acetyltransferase (Pat)/Sir2-dependent system. Taken together, this work confirms the final four steps of the 3HP/4HB pathway, thereby providing the framework for examining connections between CO2fixation and central metabolism inM. sedula.

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