Transglycosylation Activity of Engineered Bifidobacterium Lacto-N-Biosidase Mutants at Donor Subsites for Lacto-N-Tetraose Synthesis

The health benefits of human milk oligosaccharides (HMOs) make them attractive targets as supplements for infant formula milks. However, HMO synthesis is still challenging and only two HMOs have been marketed. Engineering glycoside hydrolases into transglycosylases may provide biocatalytic routes to the synthesis of complex oligosaccharides. Lacto-N-biosidase from Bifidobacterium bifidum (LnbB) is a GH20 enzyme present in the gut microbiota of breast-fed infants that hydrolyzes lacto-N-tetraose (LNT), the core structure of the most abundant type I HMOs. Here we report a mutational study in the donor subsites of the substrate binding cleft with the aim of reducing hydrolytic activity and conferring transglycosylation activity for the synthesis of LNT from p-nitrophenyl β-lacto-N-bioside and lactose. As compared with the wt enzyme with negligible transglycosylation activity, mutants with residual hydrolase activity within 0.05% to 1.6% of the wild-type enzyme result in transglycosylating enzymes with LNT yields in the range of 10–30%. Mutations of Trp394, located in subsite -1 next to the catalytic residues, have a large impact on the transglycosylation/hydrolysis ratio, with W394F being the best mutant as a biocatalyst producing LNT at 32% yield. It is the first reported transglycosylating LnbB enzyme variant, amenable to further engineering for practical enzymatic synthesis of LNT.     

Interleukin-1β Modulation of the Mechanobiology of Primary Human Pulmonary Fibroblasts: Potential Implications in Lung Repair

Pro-inflammatory cytokines like interleukin-1β (IL-1β) are upregulated during early responses to tissue damage and are expected to transiently compromise the mechanical microenvironment. Fibroblasts are key regulators of tissue mechanics in the lungs and other organs. However, the effects of IL-1β on fibroblast mechanics and functions remain unclear. Here we treated human pulmonary fibroblasts from control donors with IL-1β and used Atomic Force Microscopy to unveil that IL-1β significantly reduces the stiffness of fibroblasts concomitantly with a downregulation of filamentous actin (F-actin) and alpha-smooth muscle (α-SMA). Likewise, COL1A1 mRNA was reduced, whereas that of collagenases MMP1 and MMP2 were upregulated, favoring a reduction of type-I collagen. These mechanobiology changes were functionally associated with reduced proliferation and enhanced migration upon IL-1β stimulation, which could facilitate lung repair by drawing fibroblasts to sites of tissue damage. Our observations reveal that IL-1β may reduce local tissue rigidity by acting both intracellularly and extracellularly through the downregulation of fibroblast contractility and type I collagen deposition, respectively. These IL-1β-dependent mechanical effects may enhance lung repair further by locally increasing pulmonary tissue compliance to preserve normal lung distension and function. Moreover, our results support that IL-1β provides innate anti-fibrotic protection that may be relevant during the early stages of lung repair.     

A thermo-mechanical machining method for improving the accuracy and stability of the geometric shape of long low-rigidity shafts

Journal of Intelligent Manufacturing - The article presents a new thermo-mechanical machining method for the manufacture of long low-rigidity shafts which combines straightening and heat treatment...     

Wide-Area Crowd Counting: Multi-view Fusion Networks for Counting in Large Scenes

International Journal of Computer Vision - Crowd counting in single-view images has achieved outstanding performance on existing counting datasets. However, single-view counting is not applicable...     

Studies on the effect of humic acids and phenol on adsorption-ultrafiltration process performance

Application of pressure-driven membrane processes, such as ultrafiltration (UF) and microfiltration (MF) for surface water treatment have become very popular during last decades. Membrane fouling by humic substances (HS) is one of the major limiting factors in these processes. In order to alleviate the unfavorable effects of the presence of HS in the feed on the process performance UF and MF are often combined with adsorption on powdered activated carbon (PAC). The main goal of the present study was to evaluate the effect of humic acid (HA) on membrane fouling during UF. Moreover, the effect of PAC addition to the feed on UF process, especially on flux decline was determined. The applicability of the adsorption-ultrafiltration (PAC/UF) system to purification of water containing low (phenol) and high molecular (HA) was also investigated. Three different polymer UF membranes, prepared from polysulfone (PSF), cellulose acetate (CA) or polyacrylonitrile (PAN) were applied. It was found that the membranes prepared from PSF and CA are very susceptible to fouling caused by HA. The permeate flux decreased for ca. 50% during UF of HA solution through the PSF membrane and for ca. 45%-through the CA membrane. In the case of the PAN membrane, a negligible effect of HA on the flux was observed. On the basis of the FTIR spectra it was found that the drop in the permeate flux through these membranes may result from interactions between the negatively charged functional groups present on the membrane surface, such as carboxyl groups (CA) and sulfone groups (PSF) with HA, which results in coating of the membrane surface with HA. When PAC was added to the feed containing HA, the permeate flux through the CA and PAN membranes was maintained on a practically unchanged level. However, in case of the PSF membrane, a 50% drop in the permeate flux in comparison with the flux value, when process was conducted without PAC addition was observed. That was supposed to be due to attractive forces among hydrophobic PAC particles, HA molecules and PSF membrane surface. The performed studies showed that the application of PAC/UF system was very effective in the removal of organic substances having both, low and high molecular weights. The role of PAC suspended in a feed in the PAC/UF system is the adsorption of low molecular organic compounds, which cannot be removed by UF alone.     

Design and implementation of an ultrasonic sensor for rapid monitoring of industrial malolactic fermentation of wines

Ultrasound is an emerging technology that can be applied to monitor food processes. However, ultrasonic techniques are usually limited to research activities within a laboratory environment and they are not extensively used in industrial processes. The aim of this paper is to describe a novel ultrasonic sensor designed to monitor physical–chemical changes that occur in wines stored in industrial tanks. Essentially, the sensor consists of an ultrasonic transducer in contact with a buffer rod, mounted inside a stainless steel tube section. This structure allows the ultrasonic sensor to be directly installed in stainless steel tanks of an industrial plant. The operating principle of this design is based on the measurement of ultrasonic velocity of propagation. To test its proper operation, the sensor has been used to measure changes of concentration in aqueous samples and to monitor the progress of a malolactic fermentation of red wines in various commercial wineries. Results show the feasibility of using this sensor for monitoring malolactic fermentations in red wines placed in industrial tanks.     

3D Radiation Therapy Boost Improves the Outcome of Whole Brain Radiation Therapy Treated RPA II Patients with One or Two Brain Metastases

Purpose

to evaluate the role of whole brain radiotherapy (WBRT) and radiation boost (RB) for 208 patients recursive partitioning analysis (RPA) II with 1 or 2 brain metastases (BM) at a single institution.

Methods and Materials

the dose of WBRT was 30 Gy (10 fractions of 3 Gy). One hundred thirty-two patients (63.5%) benefited from RB of 9 Gy in 3 fractions of 3 Gy at the metastatic site. Patients had 1 or 2 BM in 122 (58.7%) and 86 cases (41.3%), respectively.

Results

patients with one or two metastases had similar survival (4.6 and 5.1 months, respectively) (p = 0.4). Median overall survival (OS) for patients treated with WBRT and RB, and with WBRT alone was 5.9 and 3.7 months, respectively (p = 0.03). The 6-, 12- and 24-month OS rates after WBRT and RB were 48.5%, 25% and 10.6%, respectively, while WBRT alone resulted in OS rates of 34%, 22.4% and 3.2%, respectively (p = 0.03). After WBRT and RB, the 6-, 12- and 24-month local control rates were 92%, 82% and 67%, respectively, while they were 81.2%, 75% and 37.5%, respectively, after WBRT alone (p = 0.03). The 6-, 12- and 24-month brain control rates after WBRT and RB were 88.7%, 75.8% and 62%, respectively, and after WBRT alone they were 78.5%, 59% and 37.7%, respectively (p = 0.03).

Conclusion

additional boost delivered with 3D conformal radiotherapy improves local and brain control rates significantly as well as overall survival for RPA II patients with 1 or 2 unresectable BM.     

Behaviour of Dietary Fibre Supplements During Bread Dough Development Evaluated Using Novel Farinograph Curve Analysis

A regression model composed of two sigmoid functions with variable asymptotes was developed and used to analyse the dietary fibre supplement behaviour during bread dough mixing. Using eight commercial dietary fibres of different botanical origin and model wheat starch-gluten flour, farinograph curves for fibre-flour blends and for the flour only were performed. After smoothing of the curves with the regression model, difference farinograms as the fibre supplementation effects were determined. The results showed that the difference farinograms had two peaks, whose shape was strongly differentiated by the studied supplements. The presence of both peaks allowed distinguishing two kinds of the rheological activity of each fibre supplement: weakening and strengthening of the consistency of bread dough during its development. The carrot, oat, cranberry, and cacao fibres exhibited dominance of strengthening over weakening action. While chokeberry, carob, apple, and flax fibres were characterised by dominance of weakening over strengthening action. The analysis of both position and height of the peaks in the difference farinogram can be also helpful for determination of hydration kinetics of the fibres used for bread supplementation.     

Microwave vacuum drying and puffing of the meat tissue - process analysis

Tenderization based on papain and microwave vacuum drying and puffing (MVDP) as well as adequate processing conditions enabled expansion of slices of the raw meat tissue. The main aim of the study was to use the response surface method and find the conditions of puffing that could maximize the expansion of meat during MVDP. The bound was set on the expansion: the most expanded puffings should be of good quality, including a desirable porous structure. The best combination of values of the MVDP process conditions (e.g., the initial moisture content in meat slices, microwave energy, and the vacuum absolute pressure) was found during experiments using the central composite design. The volume of slices puffed under these conditions was 1.60–1.78 times greater than the volume of slices before MVDP, and the sensory quality index remained acceptable. The research showed that pretreatment reduced the hardness of sample by over 25% and increased its crispiness, which was confirmed in cryo-scanning electron microscopy images.