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Frontiers in Cellular and Infection... 2022Protein lysine malonylation (Kmal) is a novel post-translational modification (PTM) that regulates various biological pathways such as energy metabolism and translation....
Protein lysine malonylation (Kmal) is a novel post-translational modification (PTM) that regulates various biological pathways such as energy metabolism and translation. Malonylation in prokaryotes, however, is still poorly understood. In this study, we performed a global Kmal analysis of the cariogenic organism by combining antibody-based affinity enrichment and high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) analysis. Altogether, 392 malonyllysine sites in 159 proteins were identified. Subsequent bioinformatic analysis revealed that Kmal occurs in proteins involved in various metabolic pathways including translation machinery, energy metabolism, RNA degradation, and biosynthesis of various secondary metabolites. Quantitative analysis demonstrated that Kmal substrates were globally altered in the biofilm growth state compared to the planktonic growth state. Furthermore, a comparative analysis of the lysine malonylome of our study with previously determined lysine acetylome in revealed that a small proportion of Kmal sites overlapped with acetylated sites, whereby suggesting that these two acylations have distinct functional implications. These results expand our knowledge of Kmal in prokaryotes, providing a resource for researching metabolic regulation of bacterial virulence and physiological functions by PTM.
Topics: Lysine; Malonates; Streptococcus mutans; Tandem Mass Spectrometry; Protein Processing, Post-Translational; Proteins; Acetylation
PubMed: 36519128
DOI: 10.3389/fcimb.2022.1078572 -
Diabetes & Metabolism Journal Nov 2021Brown adipose tissue (BAT) is a specialized tissue for nonshivering thermogenesis to dissipate energy as heat. Although BAT research has long been limited mostly in... (Review)
Review
Brown adipose tissue (BAT) is a specialized tissue for nonshivering thermogenesis to dissipate energy as heat. Although BAT research has long been limited mostly in small rodents, the rediscovery of metabolically active BAT in adult humans has dramatically promoted the translational studies on BAT in health and diseases. Moreover, several remarkable advancements have been made in brown fat biology over the past decade: The molecular and functional analyses of inducible thermogenic adipocytes (socalled beige adipocytes) arising from a developmentally different lineage from classical brown adipocytes have been accelerated. In addition to a well-established thermogenic activity of uncoupling protein 1 (UCP1), several alternative thermogenic mechanisms have been discovered, particularly in beige adipocytes. It has become clear that BAT influences other peripheral tissues and controls their functions and systemic homeostasis of energy and metabolic substrates, suggesting BAT as a metabolic regulator, other than for thermogenesis. This notion is supported by discovering that various paracrine and endocrine factors are secreted from BAT. We review the current understanding of BAT pathophysiology, particularly focusing on its role as a metabolic regulator in small rodents and also in humans.
Topics: Adipocytes, Brown; Adipose Tissue, Brown; Energy Metabolism; Thermogenesis; Uncoupling Protein 1
PubMed: 34176254
DOI: 10.4093/dmj.2020.0291 -
Journal of Bacteriology May 2023Carbon monoxide (CO) serves as a source of energy and carbon for a diverse set of microbes found in anaerobic and aerobic environments. The enzymes that bacteria and... (Review)
Review
Carbon monoxide (CO) serves as a source of energy and carbon for a diverse set of microbes found in anaerobic and aerobic environments. The enzymes that bacteria and archaea use to oxidize CO depend upon complex metallocofactors that require accessory proteins for assembly and proper function. This complexity comes at a high energetic cost and necessitates strict regulation of CO metabolic pathways in facultative CO metabolizers to ensure that gene expression occurs only when CO concentrations and redox conditions are appropriate. In this review, we examine two known heme-dependent transcription factors, CooA and RcoM, that regulate inducible CO metabolism pathways in anaerobic and aerobic microorganisms. We provide an analysis of the known physiological and genomic contexts of these sensors and employ this analysis to contextualize known biochemical properties. In addition, we describe a growing list of putative transcription factors associated with CO metabolism that potentially use cofactors other than heme to sense CO.
Topics: Transcription Factors; Carbon Monoxide; Oxidation-Reduction; Heme; Gene Expression; Bacterial Proteins
PubMed: 37154694
DOI: 10.1128/jb.00332-22 -
Redox Biology May 2020The pathogenesis of many human diseases has been attributed to the over production of reactive oxygen species (ROS), particularly superoxide (O) and hydrogen peroxide... (Review)
Review
The pathogenesis of many human diseases has been attributed to the over production of reactive oxygen species (ROS), particularly superoxide (O) and hydrogen peroxide (HO), by-products of metabolism that are generated by the premature reaction of electrons with molecular oxygen (O) before they reach complex IV of the respiratory chain. To date, there are 32 known ROS generators in mammalian cells, 16 of which reside inside mitochondria. Importantly, although these ROS are deleterious at high levels, controlled and temporary bursts in HO production is beneficial to mammalian cells. Mammalian cells use sophisticated systems to take advantage of the second messaging properties of HO. This includes controlling its availability using antioxidant systems and negative feedback loops that inhibit the genesis of ROS at sites of production. At its core, ROS production depends on fuel metabolism. Therefore, desensitizing HO signals would also require the temporary inhibition of fuel combustion and fluxes through metabolic pathways that promote ROS production. Additionally, this would also demand the diversion of fuels and nutrients into pathways that generate NADPH and other molecules required to maintain cellular redox buffering capacity. Therefore, fuel selection and metabolic flux plays an integral role in dictating the strength and duration of cellular redox signals. In the present review I provide an updated view on the function of protein S-glutathionylation, a ubiquitous redox sensitive modification involving the formation of a disulfide between the small molecular antioxidant glutathione and a cysteine residue, in the regulation of cellular metabolism on a global scale. To date, these concepts have mostly been reviewed at the level of mitochondrial bioenergetics in the contexts of health and disease. Careful examination of the literature revealed that glutathionylation is a temporary inhibitor of most metabolic pathways including glycolysis, the Krebs cycle, oxidative phosphorylation, amino acid metabolism, and fatty acid combustion, resulting in the diversion of fuels towards NADPH-producing pathways and the inhibition of ROS production. Armed with this information, I propose that protein S-glutathionylation reactions desensitize HO signals emanating from catabolic pathways using a three-pronged regulatory mechanism; 1) inhibition of metabolic flux through pathways that promote ROS production, 2) diversion of metabolites towards pathways that support antioxidant defenses, and 3) direct inhibition of ROS-generating enzymes.
Topics: Animals; Humans; Hydrogen Peroxide; Oxidation-Reduction; Protein S; Reactive Oxygen Species; Superoxides
PubMed: 32171726
DOI: 10.1016/j.redox.2020.101472 -
Plant Communications Jan 2021Metabolons are transient multi-protein complexes of sequential enzymes that mediate substrate channeling. They differ from multi-enzyme complexes in that they are... (Review)
Review
Metabolons are transient multi-protein complexes of sequential enzymes that mediate substrate channeling. They differ from multi-enzyme complexes in that they are dynamic, rather than permanent, and as such have considerably lower dissociation constants. Despite the fact that a huge number of metabolons have been suggested to exist in plants, most of these claims are erroneous as only a handful of these have been proven to channel metabolites. We believe that physical protein-protein interactions between consecutive enzymes of a pathway should rather be called enzyme-enzyme assemblies. In this review, we describe how metabolons are generally assembled by transient interactions and held together by both structural elements and non-covalent interactions. Experimental evidence for their existence comes from protein-protein interaction studies, which indicate that the enzymes physically interact, and direct substrate channeling measurements, which indicate that they functionally interact. Unfortunately, advances in cell biology and proteomics have far outstripped those in classical enzymology and flux measurements, rendering most reports reliant purely on interactome studies. Recent developments in co-fractionation mass spectrometry will likely further exacerbate this bias. Given this, only dynamic enzyme-enzyme assemblies in which both physical and functional interactions have been demonstrated should be termed metabolons. We discuss the level of evidence for the manifold plant pathways that have been postulated to contain metabolons and then list examples in both primary and secondary metabolism for which strong evidence has been provided to support these claims. In doing so, we pay particular attention to experimental and mathematical approaches to study metabolons as well as complexities that arise in attempting to follow them. Finally, we discuss perspectives for improving our understanding of these fascinating but enigmatic interactions.
Topics: Carrier Proteins; Metabolic Networks and Pathways; Multienzyme Complexes; Plant Physiological Phenomena; Plant Proteins; Secondary Metabolism
PubMed: 33511342
DOI: 10.1016/j.xplc.2020.100081 -
ELife Aug 2020Quantitative measurements of biomolecule associations are central to biological understanding and are needed to build and test predictive and mechanistic models. Given...
Quantitative measurements of biomolecule associations are central to biological understanding and are needed to build and test predictive and mechanistic models. Given the advances in high-throughput technologies and the projected increase in the availability of binding data, we found it especially timely to evaluate the current standards for performing and reporting binding measurements. A review of 100 studies revealed that in most cases essential controls for establishing the appropriate incubation time and concentration regime were not documented, making it impossible to determine measurement reliability. Moreover, several reported affinities could be concluded to be incorrect, thereby impacting biological interpretations. Given these challenges, we provide a framework for a broad range of researchers to evaluate, teach about, perform, and clearly document high-quality equilibrium binding measurements. We apply this framework and explain underlying fundamental concepts through experimental examples with the RNA-binding protein Puf4.
Topics: Case-Control Studies; Kinetics; Ligands; Protein Binding; RNA-Binding Proteins; Reproducibility of Results; Saccharomyces cerevisiae Proteins; Thermodynamics
PubMed: 32758356
DOI: 10.7554/eLife.57264 -
Nature Communications May 2021Atmospheric oxygen is thought to have played a vital role in the evolution of large, complex multicellular organisms. Challenging the prevailing theory, we show that the...
Atmospheric oxygen is thought to have played a vital role in the evolution of large, complex multicellular organisms. Challenging the prevailing theory, we show that the transition from an anaerobic to an aerobic world can strongly suppress the evolution of macroscopic multicellularity. Here we select for increased size in multicellular 'snowflake' yeast across a range of metabolically-available O levels. While yeast under anaerobic and high-O conditions evolved to be considerably larger, intermediate O constrained the evolution of large size. Through sequencing and synthetic strain construction, we confirm that this is due to O-mediated divergent selection acting on organism size. We show via mathematical modeling that our results stem from nearly universal evolutionary and biophysical trade-offs, and thus should apply broadly. These results highlight the fact that oxygen is a double-edged sword: while it provides significant metabolic advantages, selection for efficient use of this resource may paradoxically suppress the evolution of macroscopic multicellular organisms.
Topics: Aerobiosis; Anaerobiosis; Biological Evolution; Biophysical Phenomena; DNA-Binding Proteins; Directed Molecular Evolution; Eukaryotic Cells; Gene Deletion; Genetic Engineering; Models, Biological; Oxygen; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Selection, Genetic; Synthetic Biology; Transcription Factors
PubMed: 33990594
DOI: 10.1038/s41467-021-23104-0 -
FEMS Yeast Research Aug 2022In response to osmotic dehydration cells sense, signal, alter gene expression, and metabolically counterbalance osmotic differences. The main compatible solute/osmolyte... (Review)
Review
In response to osmotic dehydration cells sense, signal, alter gene expression, and metabolically counterbalance osmotic differences. The main compatible solute/osmolyte that accumulates in yeast cells is glycerol, which is produced from the glycolytic intermediate dihydroxyacetone phosphate. This review covers recent advancements in understanding mechanisms involved in sensing, signaling, cell-cycle delays, transcriptional responses as well as post-translational modifications on key proteins in osmoregulation. The protein kinase Hog1 is a key-player in many of these events, however, there is also a growing body of evidence for important Hog1-independent mechanisms playing vital roles. Several missing links in our understanding of osmoregulation will be discussed and future avenues for research proposed. The review highlights that this rather simple experimental system-salt/sorbitol and yeast-has developed into an enormously potent model system unravelling important fundamental aspects in biology.
Topics: Glycerol; Osmoregulation; Osmotic Pressure; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Water-Electrolyte Balance
PubMed: 35927716
DOI: 10.1093/femsyr/foac035 -
Molecules (Basel, Switzerland) Jun 2020Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics is a powerful tool for identifying and quantifying proteins in biological samples,... (Review)
Review
Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics is a powerful tool for identifying and quantifying proteins in biological samples, outperforming conventional antibody-based methods in many aspects. LC-MS/MS-based proteomics studies have revealed the protein abundances of many drug-metabolizing enzymes and transporters (DMETs) in tissues relevant to drug metabolism and disposition. Previous studies have consistently demonstrated marked interindividual variability in DMET protein expression, suggesting that varied DMET function is an important contributing factor for interindividual variability in pharmacokinetics (PK) and pharmacodynamics (PD) of medications. Moreover, differential DMET expression profiles were observed across different species and in vitro models. Therefore, caution must be exercised when extrapolating animal and in vitro DMET proteomics findings to humans. In recent years, DMET proteomics has been increasingly utilized for the development of physiologically based pharmacokinetic models, and DMET proteins have also been proposed as biomarkers for prediction of the PK and PD of the corresponding substrate drugs. In sum, despite the existence of many challenges in the analytical technology and data analysis methods of LC-MS/MS-based proteomics, DMET proteomics holds great potential to advance our understanding of PK behavior at the individual level and to optimize treatment regimens via the DMET protein biomarker-guided precision pharmacotherapy.
Topics: Animals; Chromatography, Liquid; Enzymes; Humans; Inactivation, Metabolic; Membrane Transport Proteins; Proteomics; Tandem Mass Spectrometry
PubMed: 32545386
DOI: 10.3390/molecules25112718 -
Communications Biology Sep 2021TOR complex 1 (TORC1) is an evolutionarily-conserved protein kinase that controls cell growth and metabolism in response to nutrients, particularly amino acids. In...
TOR complex 1 (TORC1) is an evolutionarily-conserved protein kinase that controls cell growth and metabolism in response to nutrients, particularly amino acids. In mammals, several amino acid sensors have been identified that converge on the multi-layered machinery regulating Rag GTPases to trigger TORC1 activation; however, these sensors are not conserved in many other organisms including yeast. Previously, we reported that glutamine activates yeast TORC1 via a Gtr (Rag ortholog)-independent mechanism involving the vacuolar protein Pib2, although the identity of the supposed glutamine sensor and the exact TORC1 activation mechanism remain unclear. In this study, we successfully reconstituted glutamine-responsive TORC1 activation in vitro using only purified Pib2 and TORC1. In addition, we found that glutamine specifically induced a change in the folding state of Pib2. These findings indicate that Pib2 is a glutamine sensor that directly activates TORC1, providing a new model for the metabolic control of cells.
Topics: Apoptosis Regulatory Proteins; Gene Expression Regulation, Fungal; Glutamine; Metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Transcription Factors
PubMed: 34535752
DOI: 10.1038/s42003-021-02625-w