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Summary: Mineral weathering by microorganisms is considered to occur through a succession of mechanisms based on acidification and chelation. While the role of acidification is established, the role of siderophores is difficult to disentangle from the effect of the acidification. We took advantage of the ability of strain Collimonas pratensis PMB3(1) to weather minerals but not to acidify depending on the carbon source to address the role of siderophores in mineral weathering. We identified a single non‐ribosomal peptide synthetase (NRPS) responsible for siderophore biosynthesis in the PMB3(1) genome. By combining iron‐chelating assays, targeted mutagenesis and chemical analyses (HPLC and LC‐ESI‐HRMS), we identified the siderophore produced as malleobactin X and how its production depends on the concentration of available iron. Comparison with the genome sequences of other collimonads evidenced that malleobactin production seems to be a relatively conserved functional trait, though some collimonads harboured other siderophore synthesis systems. We also revealed by comparing the wild‐type strain and its mutant impaired in the production of malleobactin that the ability to produce this siderophore is essential to allow the dissolution of hematite under non‐acidifying conditions. This study represents the first characterization of the siderophore produced by collimonads and its role in mineral weathering. [ABSTRACT FROM AUTHOR]

Some metal‐chelating peptides have antioxidant properties, with potential nutrition, health, and cosmetics applications. This study aimed to simulate their separation on immobilized metal ion affinity chromatography from their affinity constant for immobilized metal ion determined in surface plasmon resonance, both technics are based on peptide‐metal ion interactions. In our approach, first, the affinity constant of synthetic peptides was determined by surface plasmon resonance and used as input data to numerically simulate the chromatographic separation with a transport‐dispersive model based on Langmuir adsorption isotherm. Then, chromatographic separation was applied on the same peptides to determine their retention time and compare this experimental tR with the simulated tR obtained from simulation from surface plasmon resonance data. For the investigated peptides, the relative values of tR were comparable. Hence, our study demonstrated the pertinence of such numerical simulation correlating immobilized metal ion affinity chromatography and surface plasmon resonance. [ABSTRACT FROM AUTHOR]

The cell‐specific growth rate (µ) is a critical process parameter for antibody production processes performed by animal cell cultures, as it describes the cell growth and reflects the cell physiological state. When there are changes in these parameters, which are indicated by variations of µ, the synthesis and the quality of antibodies are often affected. Therefore, it is essential to monitor and control the variations of µto assure the antibody production and achieve high product quality. In this study, a novel approach for on‐line estimation of µ was developed based on the process analytical technology initiative by using an in situ dielectric spectroscopy. Critical moments, such as significant µ decreases, were successfully detected by this method, in association with changes in cell physiology as well as with an accumulation of nonglycosylated antibodies. Thus, this method was used to perform medium renewals at the appropriate time points, maintaining the values of µ close to its maximum. Using this method, we demonstrated that the physiological state of cells remained stable, the quantity and the glycosylation quality of antibodies were assured at the same time, leading to better process performances compared with the reference feed‐harvest cell cultures carried out by using off‐line nutrient measurements. The cell‐specific growth rate (μ) is a critical process parameter for antibody production by animal cell cultures, as it reflects the cell physiological state of which changes often affect synthesis and quality of antibodies. A novel approach for on‐line estimation of μ was developed by using in situ dielectric spectroscopy and used to control medium renewals during feed‐harvest cultures. This on‐line strategy allowed to maintain an appropriate cell physiological state and consequently to assure the antibody production and glycosylation quality. [ABSTRACT FROM AUTHOR]

Abstract: The presence of aminoacylase activities was investigated in a crude extract of Streptomyces ambofaciens ATCC23877. First activities catalyzing the hydrolysis of N‐α or ε‐acetyl‐L‐lysine were identified. Furthermore, the acylation of lysine and different peptides was studied and compared with results obtained with lipase B of Candida antarctica (CALB). Different regioselectivities were demonstrated for the two classes of enzymes. CALB was able to catalyze acylation only on the ε‐position whereas the crude extract from S. ambofaciens possessed the rare ability to catalyze the N‐acylation on the α‐position of the lysine or of the amino‐acid in N‐terminal position of peptides. Two genes, SAM23877_1485 and SAM23877_1734, were identified in the genome of Streptomyces ambofaciens ATCC23877 whose products show similarities with the previously identified aminoacylases from Streptomyces mobaraensis. The proteins encoded by these two genes were responsible for the major aminoacylase hydrolytic activities. Furthermore, we show that the hydrolysis of N‐α‐acetyl‐L‐lysine could be attributed to the product of SAM23877_1734 gene. [ABSTRACT FROM AUTHOR]

The glycosylation of therapeutic monoclonal antibodies (mAbs), a known critical quality attribute, is often greatly modified during the production process by animal cells. It is essential for biopharmaceutical industries to monitor and control this glycosylation. However, current glycosylation characterization techniques involve time‐ and labor‐intensive analyses, often carried out at the end of the culture when the product is already synthesized. This study proposes a novel methodology for real‐time monitoring of antibody glycosylation site occupancy using Raman spectroscopy. It was first observed in CHO cell batch culture that when low nutrient concentrations were reached, a decrease in mAb glycosylation was induced, which made it essential to rapidly detect this loss of product quality. By combining in situ Raman spectroscopy with chemometric tools, efficient prediction models were then developed for both glycosylated and nonglycosylated mAbs. By comparing variable importance in projection profiles of the prediction models, it was confirmed that Raman spectroscopy is a powerful method to distinguish extremely similar molecules, despite the high complexity of the culture medium. Finally, the Raman prediction models were used to monitor batch and feed‐harvest cultures in situ. For the first time, it was demonstrated that the concentrations of glycosylated and nonglycosylated mAbs could be successfully and simultaneously estimated in real time with high accuracy, including their sudden variations due to medium exchanges. Raman spectroscopy can thus be considered as a promising PAT tool for feedback process control dedicated to on‐line optimization of mAb quality. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:486–493, 2018 [ABSTRACT FROM AUTHOR]

The objective of this paper is on the one hand to study the effect of heat processing (30 to 130°C for 2 h) on the stability and antioxidant activity of six flavonoids (rutin, naringin, eriodictyol, mesquitol, luteolin, and luteolin 7-O glucoside), and on the other hand to establish the relation structure-activity-stability of these compounds. The dependency on temperature of the six kinetics was well described by the Arrhenius law and the main parameters provided by this model are determined and compared in this paper. Activation energies found were 107.3 kJ/mol for rutin, 100.6 for naringin, 33.3 for mesquitol, 68.2 for eriodictyol, 51.4 for luteolin, and 120 for luteolin 7-O-glucoside. The data collected showed that glycosylated flavonoids are more resistant than aglycon flavonoids to heat treatment. Moreover, it was also observed that despite the total degradation of some flavonoids, the treated solutions still have an antioxidant activity. Practical application Flavonoids are mainly consumed in processed foods. Thus, flavonoids often undergo heat processing. Knowing the thermal stability and the evolution of their antioxidant activity are particularly relevant in the field of food processing. The results of this study allow information to be collected on the relationship between heat stability-flavonoid structure and antioxidant activity. This information will be useful for the formulation of new food products or for the design of new food processes. [ABSTRACT FROM AUTHOR]

We describe the impact of two propeptides and PedC on the production yield and the potency of recombinant pediocins produced in L actococcus lactis. On the one hand, the sequences encoding the propeptides SD or LEISSTCDA were inserted between the sequence encoding the signal peptide of Usp45 and the structural gene of the mature pediocin PA-1. On the other hand, the putative thiol-disulfide oxidoreductase PedC was coexpressed with pediocin. The concentration of recombinant pediocins produced in supernatants was determined by enzyme-linked immunosorbent assay. The potency of recombinant pediocins was investigated by measuring the minimal inhibitory concentration by agar well diffusion assay. The results show that propeptides SD or LEISSTCDA lead to an improved secretion of recombinant pediocins with apparently no effect on the antibacterial potency and that PedC increases the potency of recombinant pediocin. To our knowledge, this study reveals for the first time that pediocin tolerates fusions at the N-terminal end. Furthermore, it reveals that only expressing the pediocin structural gene in a heterologous host is not sufficient to get an optimal potency and requires the accessory protein PedC. In addition, it can be speculated that PedC catalyses the correct formation of disulfide bonds in pediocin. [ABSTRACT FROM AUTHOR]

The hepatic removal of triglyceride-rich chylomicrons during the postprandial phase represents an important step towards determining the bioavailability of dietary lipids amongst the peripheral tissues. Indeed, elevated postprandial lipemia is often associated with obesity and increased risk of coronary heart disease. The milk protein, lactoferrin, has been shown to inhibit hepatic chylomicron remnant removal by the liver, resulting in increased postprandial lipemia. Despite numerous studies on potential targets for lactoferrin, the molecular mechanisms underlying the effect of lactoferrin remain unclear. We recently demonstrated that the lipolysis stimulated lipoprotein receptor ( LSR) contributes to the removal of triglyceride-rich lipoproteins during the postprandial phase. Here, we report that while lactoferrin does not have any significant effect on LSR protein levels in mouse Hepa1-6 cells, this protein colocalizes with LSR in cells but only in the presence of oleate, which is needed to obtain LSR in its active form as lipoprotein receptor. Ligand blotting using purified LSR revealed that lactoferrin binds directly to the receptor in the presence of oleate and prevents the binding of triglyceride-rich lipoproteins. Both C- and N-lobes of lactoferrin as well as a mixture of peptides derived from its hydrolysis retained the ability to bind LSR in its active form. We propose then that the elevated postprandial lipemia observed upon lactoferrin treatment in vivo is mediated in part by its direct interaction with free fatty acid activated LSR, thus preventing clearance of chylomicrons and their remnants through the LSR pathway. Structured digital abstract • Lsr and Lsr bind by comigration in gel electrophoresis (View interaction) • Lsr binds to Lf by filter binding (View interaction) • Lf and Lsr colocalize by fluorescence microscopy (View interaction) • Lsr physically interacts with VLDL by filter binding (View interaction) [ABSTRACT FROM AUTHOR]

Prior studies on membranes have reported that unsaturations affect important bilayers properties, but most of these studies have been based on single-component membranes. This report compares the properties of a phosphatidylcholine (PC) membrane enriched in long chain PUFA (LC-PUFA), extracted and purified from salmon heads ( Salmo salar), with those of other multicomponent PC membranes from egg and soybean. Detailed description of the composition, the structural parameters, the thermal behavior, the hydration, and monolayer properties are given. Hydration properties of salmon PC showed greater adsorbance of water compared to egg and soybean PC. A greater adsorption of water was also observed for salmon PC multibilayers in bulk water before demixing compared to egg and soybean PC. An interesting feature concerns the evolution of area per lipid when increasing the water content where salmon PC showed a relative constant molecular area upon hydration. Another specific feature was the ability of salmon PC monolayer to occupy smaller molecular area upon compression than do egg and soybean PC. We hypothesize that specific hydration properties at the interface and enhanced packing properties of the LC-PUFA are responsible for this particular behavior of salmon PC membrane. [ABSTRACT FROM AUTHOR]

The glycosylation of therapeutic monoclonal antibodies (mAbs), a known critical quality attribute, is often greatly modified during the production process by animal cells. It is essential for biopharmaceutical industries to monitor and control this glycosylation. However, current glycosylation characterization techniques involve time- and labor-intensive analyses, often carried out at the end of the culture when the product is already synthesized. This study proposes a novel methodology for real-time monitoring of antibody glycosylation site occupancy using Raman spectroscopy. It was first observed in CHO cell batch culture that when low nutrient concentrations were reached, a decrease in mAb glycosylation was induced, which made it essential to rapidly detect this loss of product quality. By combining in situ Raman spectroscopy with chemometric tools, efficient prediction models were then developed for both glycosylated and nonglycosylated mAbs. By comparing variable importance in projection profiles of the prediction models, it was confirmed that Raman spectroscopy is a powerful method to distinguish extremely similar molecules, despite the high complexity of the culture medium. Finally, the Raman prediction models were used to monitor batch and feed-harvest cultures in situ. For the first time, it was demonstrated that the concentrations of glycosylated and nonglycosylated mAbs could be successfully and simultaneously estimated in real time with high accuracy, including their sudden variations due to medium exchanges. Raman spectroscopy can thus be considered as a promising PAT tool for feedback process control dedicated to on-line optimization of mAb quality. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:486-493, 2018., (© 2018 American Institute of Chemical Engineers.)