Trade-offs between effectiveness and cost in bifunctional enzyme circuit with concentration robustness.

A fundamental trade-off in biological systems is whether they consume resources to perform biological functions or save resources. Bacteria need to reliably and rapidly respond to input signals by using limited cellular resources. However, excessive resource consumption will become a burden for bacteria growth. To investigate the relationship between functional effectiveness and resource cost, we study the ubiquitous bifunctional enzyme circuit, which is robust to fluctuations in protein concentration and responds quickly to signal changes. We show that trade-off relationships exist between functional effectiveness and protein cost. Expressing more proteins of the circuit increases concentration robustness and response speed but affects bacterial growth. In particular, our study reveals a general relationship between free-energy dissipation rate, response speed, and concentration robustness. The dissipation of free energy plays an important role in the concentration robustness and response speed. High robustness can only be achieved with a large amount of free-energy consumption and protein cost. In addition, the noise of the output increases with increasing protein cost, while the noise of the response time decreases with increasing protein cost. We also calculate the trade-off relationships in the EnvZ-OmpR system and the nitrogen assimilation system, which both have the bifunctional enzyme. Similar results indicate that these relationships are mainly derived from the specific feature of the bifunctional enzyme circuits and are not relevant to the details of the models. According to the trade-off relationships, bacteria take a compromise solution that reliably performs biological functions at a reasonable cost.

Quantitative Connection between Cell Size and Growth Rate by Phospholipid Metabolism.

The processes involved in cell growth are extremely complicated even for a single cell organism such as Escherichia coli, while the relationship between growth rate and cell size is simple. We aimed to reveal the systematic link between them from the aspect of the genome-scale metabolic network. Since the growth rate reflects metabolic rates of bacteria and the cell size relates to phospholipid synthesis, a part of bacterial metabolic networks, we calculated the cell length from the cardiolipin synthesis rate, where the cardiolipin synthesis reaction is able to represent the phospholipid metabolism of Escherichia coli in the exponential growth phase. Combined with the flux balance analysis, it enables us to predict cell length and to examine the quantitative relationship between cell length and growth rate. By simulating bacteria growing in various nutrient media with the flux balance analysis and calculating the corresponding cell length, we found that the increase of the synthesis rate of phospholipid, the cell width, and the protein fraction in membranes caused the increase of cell length with growth rate. Different tendencies of phospholipid synthesis rate changing with growth rate result in different relationships between cell length and growth rate. The effects of gene deletions on cell size and growth rate are also examined. Knocking out the genes, such as Δ tktA, Δ tktB, Δ yqaB, Δ pgm, and Δ cysQ, affects growth rate largely while affecting cell length slightly. Results of this method are in good agreement with experiments.

Paenibacillus ottowii sp. nov. isolated from a fermentation system processing bovine manure.

Strain MS2379T was isolated from a pasteurized solution sample from a predominantly anaerobic fermentation system processing bovine manure in Pilot Point, Texas. Phylogenetic analyses based on both 16S rRNA gene and rpoB gene sequences showed that MS2379T was most closely related to Paenibacillus polymyxa (DSM 36T), P. jamilae (DSM 13815T), and P. peoriae (DSM 8320T), yet DNA-DNA relatedness through DNA-DNA hybridization revealed only 22.6, 32.0 and 24.7 % relatedness to these three species respectively. Rod-shaped cells of strain MS2379T are Gram-stain variable with sub-terminal, ellipsoidal, deforming endospores. The peptidoglycan contains meso-diaminopimelic acid (mDAP) and the predominant fatty acids are anteiso-C15 : 0 (61.9 %) and anteiso-C17 : 0 (11.6 %), confirming that strain MS2379T has diagnostic features of other Paenibacillus species. The G+C content of MS2379T is 45.9 mol%. Fermentation of glucose yields acid and gas end-products. The polar lipids found were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, and glycolipids, but also included some unidentified lipids, aminolipids, aminoglycolipid, and phosphatidylmethylethanolamine. The growth range of MS2379T was observed from 10-45 °C with optimal growth temperature at 30 °C. Growth was observed between pH 6-10 and up to 3 % NaCl. Unlike the most closely related Paenibacillus species, strain MS2379T was negative in the Voges-Proskauer reaction. Nucleic acid, chemotaxonomic and biochemical features support the distinctiveness of strain MS2379T. Thus, strain MS2379T represents a novel species of the genus Paenibacillus for which the name Paenibacillus ottowii sp. nov. is proposed with the type strain MS2379T (=DSM 107750T=ATCC TSD-165T).

Quantitative determination and pharmacokinetic study of fusaricidin A in mice plasma and tissues using ultra-high performance liquid chromatography-tandem mass spectrometry.

Fusaricidins are a family of cyclic lipodepsipeptides that convey antifungal and antibacterial activity. Fusaricidin A (FA) is one of the Fusaricidins major compounds and it is showing promising activity against fungi and bacteria. In the present study, a fast and sensitive ultra-high performance liquid chromatography coupled to triple quadrupole mass spectrometry (UHPLC-MS/MS) method was developed for the analysis of FA in mice plasma, liver, kidney and brain tissues. The instrument was operated in positive electrospray ionization mode. Multiple reaction monitoring (MRM) mode was performed with ion pairs of m/z: 883.5→256.3, 883.5→197.2 and 883.5→72.1 for FA. The method was validated for linearity, repeatability, accuracy, stability, limits of detection (LOD) and limits of quantification (LOQ). The LOD and LOQ were 0.01 and 0.05 ng/mL for plasma and tissues, respectively. The calibration curve (10-200 ng/mL) was linear ( r2 = 0.99). Precision and accuracy values were found to be < 10% (within acceptable limit). The pharmacokinetic and tissue distribution characteristics of FA were determined in plasma, liver, kidney and brain of CD1 mice after I.V. administration of a single dose of 15 mg/kg body weight. Highest plasma concentration (Cmax) was calculated to be 4169.97 ± 50 ng/mL with a tmax of 0.08 h. The plasma clearance rate of FA was 397.6 ± 203 mL/h with a t1/2 of 2.2 ± 0.5 h and apparent volume of distribution during the terminal phase (Vz) of 979.2 ± 318 mL. The highest tissue concentration (Cmax) was found in the liver (219 ± 14 ng/mg) at a tmax of 0.08 h followed by the kidneys (38.6 ± 16 ng/mg) at tmax of 0.2 h. FA was poorly distributed to the brain with a Cmax of 0.45 ± 0.2 ng/mg and a tmax of 0.08 h. The method for quantitative analysis and pharmacokinetic data provided will support the development of various formulation approaches and therapeutic application for future clinical studies.

Identification of fusaricidins from the antifungal microbial strain Paenibacillus sp. MS2379 using ultra-high performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry.

Paenibacillus sp. MS2379 is a highly efficient microbial strain producing fusaricidins, a class of lipopeptides that have demonstrated strong antifungal activities against a broad array of fungal pathogens. An integrated approach combining chromatographic fractionation, UHPLC-QTOF-MS analysis, and NMR spectroscopic interpretation was employed to characterize antifungal metabolites produced by this microbial strain, resulting in the identification of 48 fusaricidins including 30 cyclic and 18 open-chain species. In this regard, UHPLC-QTOF-MS played a vital role in determining structures of 28 new fusaricidins through peptide fragment analysis. The structural determination of the new fusaricidins by the high-resolution mass spectrometry was validated by follow-up isolation and NMR spectroscopic analysis of representative compounds. It is worth noting that novel fusaricidins with amino acid residues of serine and γ-aminobutyric acid were identified, which is of great biosynthetic significance for this biologically important class of compounds. The present study again illustrates the power of UHPLC-QTOF-MS for structural identification of lipopeptides, and the structural diversity of the identified fusaricidins makes this microbial strain unique as a potential biocontrol agent.