scholarly journals Protein interaction patterns in different cellular environments are revealed by in-cell NMR

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Letizia Barbieri ◽  
Enrico Luchinat ◽  
Lucia Banci
Keyword(s):  
Soluble Proteins ◽  
Signal Recovery ◽  
Test Case ◽  
Biological Macromolecules ◽  
Specific Interactions ◽  
Bacterial Cells ◽  
Interaction Patterns ◽  
Relative Contribution ◽  
Cellular Environment ◽  
Nmr Signals

Abstract In-cell NMR allows obtaining atomic-level information on biological macromolecules in their physiological environment. Soluble proteins may interact with the cellular environment in different ways: either specifically, with their functional partners, or non-specifically, with other cellular components. Such behaviour often causes the disappearance of the NMR signals. Here we show that by introducing mutations on the human protein profilin 1, used here as a test case, the in-cell NMR signals can be recovered. In human cells both specific and non-specific interactions are present, while in bacterial cells only the effect of non-specific interactions is observed. By comparing the NMR signal recovery pattern in human and bacterial cells, the relative contribution of each type of interaction can be assessed. This strategy allows detecting solution in-cell NMR spectra of soluble proteins without altering their fold, thus extending the applicability of in-cell NMR to a wider range of proteins.

scholarly journals Assessing the Contribution of Seasonality, Tides, and Microbial Processing to Dissolved Organic Matter Composition Variability in a Southeastern U.S. Estuary

2021 ◽  
Vol 8 ◽  
Author(s):  
Rachel P. Martineac ◽  
Alexey V. Vorobev ◽  
Mary Ann Moran ◽  
Patricia M. Medeiros
Keyword(s):  
Organic Matter ◽  
Dissolved Organic Matter ◽  
Critical Role ◽  
Bacterial Cells ◽  
Secondary Importance ◽  
Relative Contribution ◽  
Doc Concentration ◽  
Compositional Changes ◽  
Dom Composition

Uncovering which biogeochemical processes have a critical role controlling dissolved organic matter (DOM) compositional changes in complex estuarine environments remains a challenge. In this context, the aim of this study is to characterize the dominant patterns of variability modifying the DOM composition in an estuary off the Southeastern U.S. We collected water samples during three seasons (July and October 2014 and April 2015) at both high and low tides and conducted short- (1 day) and long-term (60 days) dark incubations. Samples were analyzed for bulk DOC concentration, and optical (CDOM) and molecular (FT-ICR MS) compositions and bacterial cells were collected for metatranscriptomics. Results show that the dominant pattern of variability in DOM composition occurs at seasonal scales, likely associated with the seasonality of river discharge. After seasonal variations, long-term biodegradation was found to be comparatively more important in the fall, while tidal variability was the second most important factor correlated to DOM composition in spring, when the freshwater content in the estuary was high. Over shorter time scales, however, the influence of microbial processing was small. Microbial data revealed a similar pattern, with variability in gene expression occurring primarily at the seasonal scale and tidal influence being of secondary importance. Our analyses suggest that future changes in the seasonal delivery of freshwater to this system have the potential to significantly impact DOM composition. Changes in residence time may also be important, helping control the relative contribution of tides and long-term biodegradation to DOM compositional changes in the estuary.

Formation of Two-Dimensional Crystals of Membrane-Anchored and Water-Soluble Proteins

1993 ◽  
Vol 330 ◽  
Author(s):  
Harald Engelhardt ◽  
T. Scheybani ◽  
W. Von Gustedt ◽  
W. Baumeister
Keyword(s):  
Molecular Sieves ◽  
Soluble Proteins ◽  
Water Soluble ◽  
Biological Macromolecules ◽  
Complex Structures ◽  
Two Dimensional ◽  
Protein Species ◽  
Future Developments ◽  
Multiple Protein ◽  
Support Devices

ABSTRACTThe formation of two-dimensional (2-D) crystals of biological macromolecules is of interest for nanotechnological applications. Protein 2-D crystals may be used as molecular sieves and/or support devices as components of biosensors etc. [1]. Functionally specialized 2-D crystals, containing transport or catalytic proteins, provide a certain function in a highly efficient and vectorial manner. Future developments may allow the design of more complex structures such as multilayers made from different proteins or arrays of functionally linked oligo- or multimeric complexes consisting of multiple protein species [2]. Regular 2-D arrays, either truely crystalline or densely packed molecules, are one of the basic structures taht might be used to construct more sophisticated protein-based devices. 2-D crystals have some interesting features:

scholarly journals Non-specific interactions between soluble proteins and lipids induce irreversible changes in the properties of lipid bilayers

Soft Matter ◽  
2013 ◽  
Vol 9 (16) ◽  
pp. 4219-4226 ◽  
Author(s):  
Francesca Ruggeri ◽  
Fan Zhang ◽  
Tania Lind ◽  
Erica D. Bruce ◽  
Boris L. T. Lau ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Soluble Proteins ◽  
Specific Interactions

scholarly journals Charge-driven condensation of RNA and proteins suggests broad role of phase separation in cytoplasmic environments

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Bercem Dutagaci ◽  
Grzegorz Nawrocki ◽  
Joyce Goodluck ◽  
Ali Akbar Ashkarran ◽  
Charles G Hoogstraten ◽  
...  
Keyword(s):  
Phase Separation ◽  
Molecular Size ◽  
Biological Macromolecules ◽  
Separation Processes ◽  
Cellular Environment ◽  
Polymer Properties ◽  
Protein Components ◽  
Separation Of Mixtures ◽  
Broad Role

Phase separation processes are increasingly being recognized as important organizing mechanisms of biological macromolecules in cellular environments. Well-established drivers of phase separation are multi-valency and intrinsic disorder. Here, we show that globular macromolecules may condense simply based on electrostatic complementarity. More specifically, phase separation of mixtures between RNA and positively charged proteins is described from a combination of multiscale computer simulations with microscopy and spectroscopy experiments. Phase diagrams were mapped out as a function of molecular concentrations in experiment and as a function of molecular size and temperature via simulations. The resulting condensates were found to retain at least some degree of internal dynamics varying as a function of the molecular composition. The results suggest a more general principle for phase separation that is based primarily on electrostatic complementarity without invoking polymer properties as in most previous studies. Simulation results furthermore suggest that such phase separation may occur widely in heterogenous cellular environment between nucleic acid and protein components.

scholarly journals Bacteria autoaggregation: how and why bacteria stick together

2021 ◽  
Author(s):  
El-shama Q. A. Nwoko ◽  
Iruka N. Okeke
Keyword(s):  
Protein Interaction ◽  
Biofilm Formation ◽  
Ex Vivo ◽  
Therapeutic Targets ◽  
Phase Variation ◽  
Bacterial Adherence ◽  
Specific Interactions ◽  
Bacterial Cells ◽  
Qualitative And Quantitative

Autoaggregation, adherence between identical bacterial cells, is important for colonization, kin and kind recognition, and survival of bacteria. It is directly mediated by specific interactions between proteins or organelles on the surfaces of interacting cells or indirectly by the presence of secreted macromolecules such as eDNA and exopolysaccharides. Some autoaggregation effectors are self-associating and present interesting paradigms for protein interaction. Autoaggregation can be beneficial or deleterious at specific times and niches. It is, therefore, typically regulated through transcriptional or post-transcriptional mechanisms or epigenetically by phase variation. Autoaggregation can contribute to bacterial adherence, biofilm formation or other higher-level functions. However, autoaggregation is only required for these phenotypes in some bacteria. Thus, autoaggregation should be detected, studied and measured independently using both qualitative and quantitative in vitro and ex vivo methods. If better understood, autoaggregation holds the potential for the discovery of new therapeutic targets that could be cost-effectively exploited.

scholarly journals Charge-Driven Condensation of RNA and Proteins Suggests Broad Role of Phase Separation in Cytoplasmic Environments

2020 ◽  
Author(s):  
Bercem Dutagaci ◽  
Grzegorz Nawrocki ◽  
Joyce Goodluck ◽  
Ali Akbar Ashkarran ◽  
Charles G. Hoogstraten ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Molecular Size ◽  
Liquid Phase Separation ◽  
Coarse Grained ◽  
Biological Macromolecules ◽  
Separation Processes ◽  
Cellular Environment ◽  
Liquid Liquid Phase Separation

ABSTRACTPhase separation processes are increasingly being recognized as important organizing mechanisms of biological macromolecules in cellular environments. Well established drivers of liquid-liquid phase separation are multi-valency and intrinsic disorder. Here, we show that globular macromolecules may condense simply based on electrostatic complementarity. More specifically, phase separation of mixtures between RNA and positively charged proteins is described from a combination of multiscale computer simulations with microscopy and spectroscopy experiments. Condensates retain liquid character and phase diagrams are mapped out as a function of molecular concentrations in experiment and as a function of molecular size and temperature via simulations. The results suggest a more general principle for phase separation that is based primarily on electrostatic complementarity without invoking polymer properties as in most previous studies. Simulation results furthermore suggest that such phase separation may occur widely in heterogenous cellular environment between nucleic acid and protein components.STATEMENT OF SIGNIFICANCELiquid-liquid phase separation has been recognized as a key mechanism for forming membrane-less organelles in cells. Commonly discussed mechanisms invoke a role of disordered peptides and specific multi-valent interactions. We report here phase separation of RNA and proteins based on a more universal principle of charge complementarity that does not require disorder or specific interactions. The findings are supported by coarse-grained simulations, theory, and experimental validation via microscopy and spectroscopy. The broad implication of this work is that condensate formation may be a universal phenomenon in biological systems.

scholarly journals Both galactosaminogalactan and α-1,3-glucan contribute to aggregation of Aspergillus oryzae hyphae in liquid culture

2019 ◽  
Author(s):  
Ken Miyazawa ◽  
Akira Yoshimi ◽  
Motoaki Sano ◽  
Fuka Tabata ◽  
Asumi Sugahara ◽  
...  
Keyword(s):  
Filamentous Fungi ◽  
Aspergillus Oryzae ◽  
Double Mutant ◽  
Liquid Culture ◽  
Bacterial Cells ◽  
Wild Type ◽  
Hydrogen Bond Formation ◽  
Relative Contribution ◽  
In The Wild ◽  
Increase Productivity

AbstractFilamentous fungi generally form aggregated hyphal pellets in liquid culture. We previously reported that α-1,3-glucan-deficient mutants of Aspergillus nidulans did not form hyphal pellets and their hyphae were fully dispersed, and we suggested that α-1,3-glucan functions in hyphal aggregation. Yet, Aspergillus oryzae α-1,3-glucan-deficient (AGΔ) mutants still form small pellets; therefore, we hypothesized that another factor responsible for forming hyphal pellets remains in these mutants. Here, we identified an extracellular matrix polysaccharide galactosaminogalactan (GAG) as such a factor. To produce a double mutant of A. oryzae (AG-GAGΔ), we disrupted the genes required for GAG biosynthesis in an AGΔ mutant. Hyphae of the double mutant were fully dispersed in liquid culture, suggesting that GAG is involved in hyphal aggregation in A. oryzae. Addition of partially purified GAG fraction to the hyphae of the AG-GAGΔ strain resulted in formation of mycelial pellets. Acetylation of the amino group in galactosamine of GAG weakened GAG aggregation, suggesting that hydrogen bond formation by this group is important for aggregation. Genome sequences suggest that α-1,3-glucan, GAG, or both are present in many filamentous fungi and thus may function in hyphal aggregation in these fungi. We also demonstrated that production of a recombinant polyesterase, CutL1, was higher in the AG-GAGΔ strain than in the wild-type and AGΔ strains. Thus, controlling hyphal aggregation factors of filamentous fungi may increase productivity in the fermentation industry.ImportanceProduction using filamentous fungi is an important part of the fermentation industry, but hyphal aggregation in these fungi in liquid culture limits productivity compared with that of yeast or bacterial cells. We found that galactosaminogalactan and α-1,3-glucan both function in hyphal aggregation in Aspergillus oryzae, and that the hyphae of a double mutant deficient in both polysaccharides become fully dispersed in liquid culture. We also revealed the relative contribution of α-1,3-glucan and galactosaminogalactan to hyphal aggregation. Recombinant protein production was higher in the double mutant than in the wild-type strain. Our research provides a potential technical innovation for the fermentation industry that uses filamentous fungi, as regulation of the growth characteristics of A. oryzae in liquid culture may increase productivity.

scholarly journals Expanded Genomic Sampling of the Desulfobulbales Reveals Distribution and Evolution of Sulfur Metabolisms

2021 ◽  
Author(s):  
Lewis M. Ward ◽  
Emma Bertran ◽  
David T. Johnston
Keyword(s):  
Sulfate Reduction ◽  
Sulfur Metabolism ◽  
Phylogenetic Analyses ◽  
Genomic Diversity ◽  
Test Case ◽  
Sulfur Disproportionation ◽  
Sulfate Reducing ◽  
Relative Contribution ◽  
Microbial Sulfate Reduction ◽  
Reducing Bacteria

AbstractThe reconstruction of modern and paleo-sulfur cycling relies on understanding the long-term relative contribution of its main actors; these include microbial sulfate reduction (MSR) and microbial sulfur disproportionation (MSD). However, a unifying theory is lacking for how MSR and MSD, with the same enzyme machinery and intimately linked evolutionary histories, perform two drastically different metabolisms. Here, we aim at shedding some light on the distribution, diversity, and evolutionary histories of MSR and MSD, with a focus on the Desulfobulbales as a test case. The Desulfobulbales is a diverse and widespread order of bacteria in the Desulfobacterota (formerly Deltaproteobacteria) phylum primarily composed of sulfate reducing bacteria. Recent culture- and sequence-based approaches have revealed an expanded diversity of organisms and metabolisms within this clade, including the presence of obligate and facultative sulfur disproportionators. Here, we present draft genomes of previously unsequenced species of Desulfobulbales, substantially expanding the available genomic diversity of this clade. We leverage this expanded genomic sampling to perform phylogenetic analyses, revealing an evolutionary history defined by vertical inheritance of sulfur metabolism genes with numerous convergent instances of transition from sulfate reduction to sulfur disproportionation.

scholarly journals Interaction of Pneumococcal Histidine Triad Proteins with Human Complement

2010 ◽  
Vol 78 (5) ◽  
pp. 2089-2098 ◽  
Author(s):  
Merit Melin ◽  
Emmanuel Di Paolo ◽  
Leena Tikkanen ◽  
Hanna Jarva ◽  
Cecile Neyt ◽  
...  
Keyword(s):  
Factor H ◽  
Surface Proteins ◽  
Bacterial Cells ◽  
Wild Type ◽  
Protein Antigens ◽  
Pneumococcal Infections ◽  
Relative Contribution ◽  
Bacterial Lysates ◽  
Affect Factor ◽  
Complement Deposition

ABSTRACT The pneumococcal histidine triad (Pht) proteins PhtA, PhtB, PhtD, and PhtE form a group of conserved pneumococcal surface proteins. Humans produce antibodies to Pht proteins upon exposure to pneumococcus, and immunization of mice has provided protective immunity against sepsis and pneumonia and reduced nasopharyngeal colonization. Pht proteins are candidates for inclusion in multicomponent pneumococcal protein vaccines. Their biological function in pneumococcal infections is not clear, but a role in complement inhibition has been suggested. We measured complement deposition on wild-type and Pht mutant strains in four genetic backgrounds: Streptococcus pneumoniae D39 (serotype 2) and R36A (unencapsulated derivative of D39) and strains of serotypes 3, 4, and 19F. PspA and PspC single and double mutants were compared to the wild-type and Pht-deficient D39 strains. Factor H binding was measured to bacterial cells, lysates, and protein antigens. Deletion of all four Pht proteins (Pht−) resulted in increased C3 deposition on the serotype 4 strain but not on the other strains. Pht antigens did not bind factor H, and deletion of Pht proteins did not affect factor H binding by bacterial lysates. The Pht− mutant serotype 4 strain bound slightly less factor H than the wild-type strain when binding was measured by flow cytometry. Pht proteins may play a role in immune evasion, but the mechanism of function is unlikely to be mediated by factor H binding. The relative contribution of Pht proteins to the inhibition of complement deposition is likely to be affected by the presence of other pneumococcal proteins and to depend on the genetic background.

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