Analysis of Transglucosylation Products of Aspergillus niger α-Glucosidase that Catalyzes the Formation of α-1,2- and α-1,3-Linked Oligosaccharides

According to whole-genome sequencing, Aspergillus niger produces multiple enzymes of glycoside hydrolases (GH) 31. Here we focus on a GH31 α-glucosidase, AgdB, from A. niger . AgdB has also previously been reported as being expressed in the yeast species, Pichia pastoris ; while the recombinant enzyme (rAgdB) has been shown to catalyze tranglycosylation via a complex mechanism. We constructed an expression system for A. niger AgdB using Aspergillus nidulans . To better elucidate the complicated mechanism employed by AgdB for transglucosylation, we also established a method to quantify glucosidic linkages in the transglucosylation products using 2D NMR spectroscopy. Results from the enzyme activity analysis indicated that the optimum temperature was 65 °C and optimum pH range was 6.0–7.0. Further, the NMR results showed that when maltose or maltopentaose served as the substrate, α-1,2-, α-1,3-, and small amount of α-1,1-β-linked oligosaccharides are present throughout the transglucosylation products of AgdB. These results suggest that AgdB is an α-glucosidase that serves as a transglucosylase capable of effectively producing oligosaccharides with α-1,2-, α-1,3-glucosidic linkages.     

Delamination growth mechanisms in woven glass fiber reinforced polymer composites under Mode II fatigue loading at cryogenic temperatures

The purpose of this research is to characterize the cryogenic delamination growth behavior in woven glass fiber reinforced polymer (GFRP) composite laminates subjected to Mode II fatigue loading. Mode II fatigue delamination tests were performed at room temperature, liquid nitrogen temperature (77 K) and liquid helium temperature (4 K) using the four-point bend end-notched flexure (4ENF) test method, and the delamination growth rate data for the woven GFRP laminates were obtained. The energy release rate range was determined by the finite element method. Microscopic examinations of the specimen sections and fracture surfaces were also carried out. The present results are discussed to obtain an understanding of the fatigue delamination growth mechanisms in the woven GFRP laminates under Mode II loading at cryogenic temperatures.     

The ({beta}{alpha})8 glycosidases: sequence and structure analyses suggest distant evolutionary relationships

There are currently at least nine distinct glycosidase sequencefamilies which are all known to adopt a TIM barrel fold [Henrissat,B.and Davies,G. (1997) Curr. Opin. Struct. Biol., 7, 637–644].To explore the relationships between these enzymes and theirevolution, comprehensive sequence and structure comparisonswere performed, generating four distinct clusters. The firstcluster, S1, comprises the     

Analytic Flow Design Method for an Automated Distribution Center with Multiple Shipping Areas

An analytic flow design algorithm for an automated distribution center with multiple shipping areas is presented. The main elements of the analytic flow design are the number of devices and the volumes of product flow between the devices. In the design of an automated distribution center, it is necessary to satisfy the demand throughput while minimizing construction costs. In the conventional design process, system engineers utilize experimental and intuitional approaches. However, conventional approaches are time-consuming and the design outcome is dependent on the skill of the designer. Therefore, a theoretical design algorithm is needed. We propose an analytic flow design algorithm using a dynamic network flow model and considering time-variable flow volumes according to shipment and storage schedules. To verify the feasibility of the proposed method, we perform analytic flow design using real data and confirm that the proposed method can yield a feasible analytic flow design in several minutes.     

A New Fluorogenic Probe for the Detection of endo‐β‐N‐Acetylglucosaminidase

We developed a fluorescence‐quenching‐based assay system to determine the hydrolysis activity of endo‐β‐N‐acetylglucosaminidases (ENGases). The pentasaccharide derivative 1 was labeled with an N‐methylanthraniloyl group as a reporter dye at the non‐reducing end and with a 2,4‐dinitrophenyl group as a quencher molecule at the reducing end. This derivative is hydrolyzed by ENGase, resulting in an increase in fluorescence intensity. Thus, the fluorescence signal is directly proportional to the amount of the tetrasaccharide derivative, hence allowing ENGase activity to be evaluated easily and quantitatively. Using this system, we succeeded in measuring the hydrolysis activities of ENGases and thus the inhibitory activities of known inhibitors. We confirmed that this assay system is suitable for high‐throughput screening for potential inhibitors of human ENGase that might serve as therapeutic agents for the treatment of N‐glycanase 1 (NGLY1) deficiency.     

Competitive sorption of water vapor and CO2 in photocrosslinked PVCA film for a capacitive‐type humidity sensor

The sorption behavior of water vapor and CO₂ gas in photocrosslinked poly(vinyl cinnamate) (PVCA) film was examined at 30°C under atmospheric pressure. Both the water sorption isotherm and the CO₂ sorption isotherm obtained with quartz crystal microbalance (QCM) method obeyed the simple Langmuir's equation. Water vapor/CO₂ mixed‐gas sorption isotherms were also obtained. Total amount of sorbed mixed gases was clearly influenced by the partial pressure of water vapor (p_w) and CO₂ gas (p_c) in the atmosphere. A modified Langmuir's equation based on a dual‐site model was employed for predicting the competitive adsorption isotherm, and the isotherm was clearly described by the equation. The theoretically estimated amount of adsorbed water at the constant p_w decreased slightly with increasing p_c. The effect of this phenomenon on the sensitivity of the capacitive‐type relative humidity sensor was examined. As expected, the electrical capacitance of the sensor at the constant relative humidity decreased because of the coexistence of CO₂ gas. However, the influence was quite small in the CO₂ concentration range in the ordinary environment. © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 83: 401–407, 2002     

Distribution and heterogeneity of mast cells in the human uterus

Mast cells (MCs) are widely distributed in most human tissues. Those cells that contain only tryptase are designated as T-MCs, while those that also contain chymase are referred to as TC-MCs. This study uses immunohistochemical staining for tryptase and chymase to assess the distribution and heterogeneity of these two types of MCs in the human uterus. The greatest number of MCs was found in the inner (i.e. luminal) half of the myometrium, with this area containing approximately equal proportions of T-MCs and TC-MCs. There were fewer MCs in the outer half of the myometrium and the cervix, but the proportion of TC-MCs in both of these areas was substantially higher. In contrast, the endometrium contained significantly fewer MCs, but proportionally more T-MCs. There was no change in the number of MCs between the proliferative and secretory phases of the menstrual cycle; however, there was a significantly lower number in all areas after menopause. Most of the MCs were observed in close association with uterine smooth muscle cells, as well as in the vicinity of fibroblasts and collagen, and it appears they may play an important role in the reconstruction of uterine tissues during the menstrual cycle.     

Human melanocytes and keratinocytes exposed to UVB or UVA in vivo show comparable levels of thymine dimers

Epidemiology shows a relationship between solar exposure and all types of skin cancer. Understanding the mechanisms of skin cancer requires knowledge of the photomolecular events that occur within the relevant epidermal cell types in vivo. Studies to date have focused on UVR-induced DNA lesions in keratinocytes, the majority epidermal cell population which gives rise to most skin cancers. Malignant melanoma, arising from melanocytes (5%-10% of epidermal cells), accounts for most skin cancer deaths. We report on new techniques to detect DNA photolesions in human epidermal melanocytes in situ. Previously nonexposed buttock skin of volunteers of skin types I/II was exposed to clinically relevant doses of narrow bandwidth UVB (300 nm) and UVA (320 nm, 340 nm, 360 nm) radiation. Biopsies were taken immediately afterwards and processed for routine histology. Microscope sections were prepared and double-stained with fluorescent-tagged monoclonal antibodies for thymine dimers and melanocytes. UVR dose-response curves for dimer levels within melanocyte nuclei were determined by image analysis and compared with dimer levels in adjacent basal cell keratinocytes. Our data show that UVB and UVA readily induce thymine dimers in melanocytes at levels that are comparable with those found in adjacent keratinocytes. This new technique will enable melanocyte specific studies, such as DNA repair kinetics, to be done in vivo.