Purification and characterization of an alpha1,2,-L-fucosyltransferase, which modifies the cytosolic protein FP21,from the cytosol of Dictyostelium

A novel fucosyltransferase (cFTase) activity has been enriched over 10(6)-fold from the cytosolic compartment of Dictyostelium based on transfer of [3H]fucose from GDP-[3H]fucose to Galbeta1,3 GlcNAc beta-paranitrophenyl (paranitrophenyl-lacto-N-bioside or pNP-LNB). The activity behaved as a single component during purification over DEAE-, phenyl-, Reactive Blue-4-, GDP-adipate-, GDP-hexanolamine-, and Superdex gel filtration resins. The purified activity possessed an apparent Mr of 95 X 10(3), was Mg2+-dependent with a neutral pH optimum, and exhibited a Km for GDP-fucose of 0.34 microM, a Km for pNP-LNB of 0.6 mM, and a Vmax for pN-P-LNB of 620 nmol/min/mg protein. SDS-polyacrylamide gel electrophoresis analysis of the Superdex elution profile identified a polypeptide with an apparent Mr of 85 X 10(3), which coeluted with the cFTase activity and could be specifically photolabeled with the donor substrate inhibitor GDP-hexanolaminyl-azido-125I-salicylate. Based on substrate analogue studies, exoglycosidase digestions, and co-chromatography with fucosylated standards, the product of the reaction with pNP-LNB was Fucalpha1, 2Galbeta1,3GIcNAcbeta-pNP. The cFTase preferred substrates with a Galbeta1,3linkage, and thus its acceptor substrate specificity resembles the human Secretor-type alpha1,2- FTase. Afucosyl isoforms of the FP21 glycoprotein, GP21-I and GP21-II, were purified from the cytosol of a Dictyostelium mutant and found to be substrates for the cFTase, which exhibited an apparent K(m) of 0.21 microM and an apparent V(max) of 460 nmol/min/mg protein toward GP21-II. The highly purified cFTase was inhibited by the reaction products Fucalpha1,2Galbeta1,3GlcNAcbeta-pNP and FP21-II. FP21-I and recombinant FP21 were not inhibitory, suggesting that acceptor substrate specificity is based primarily on carbohydrate recognition. A cytosolic location for this step of FP21 glycosylation is implied by the isolation of the cFTase from the cytosolic fraction, its high affinity for its substrates, and its failure to be detected in crude membrane preparations.     

Performance analysis of a COFDM/FM in-band digital audiobroadcasting system

In this paper, we present a performance analysis of a coded orthogonal frequency division multiplexing DAB system that is power multiplexed over a commercial FM station. We model the mobile channel with a time-frequency scattering function. Results show that the performance of the FM demodulator is crucial, since the level of the residual FM signal has a strong influence on the BER performance of the COFDM system. As with other mobile communication systems, the use of interleaving greatly enhances the performances     

Soluble Prokaryotic Overexpression and Purification of Human GM-CSF Using the Protein Disulfide Isomerase b′a′ Domain

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a member of the colony-stimulating factor (CSF) family, which functions to enhance the proliferation and differentiation of hematopoietic stem cells and other hematopoietic lineages such as neutrophils, dendritic cells, or macrophages. These proteins have thus generated considerable interest in clinical therapy research. A current obstacle to the prokaryotic production of human GM-CSF (hGM-CSF) is its low solubility when overexpressed and subsequent complex refolding processes. In our present study, the solubility of hGM-CSF was examined when combined with three N-terminal fusion tags in five E. coli strains at three different expression temperatures. In the five E. coli strains BL21 (DE3), ClearColi BL21 (DE3), LOBSTR, SHuffle T7 and Origami2 (DE3), the hexahistidine-tagged hGM-CSF showed the best expression but was insoluble in all cases at each examined temperature. Tagging with the maltose-binding protein (MBP) and the b′a′ domain of protein disulfide isomerase (PDIb′a′) greatly improved the soluble overexpression of hGM-CSF at 30 °C and 18 °C. The solubility was not improved using the Origami2 (DE3) and SHuffle T7 strains that have been engineered for disulfide bond formation. Two conventional chromatographic steps were used to purify hGM-CSF from the overexpressed PDIb′a′-hGM-CSF produced in ClearColi BL21 (DE3). In the experiment, 0.65 mg of hGM-CSF was isolated from a 0.5 L flask culture of these E. coli and showed a 98% purity by SDS-PAGE analysis and silver staining. The bioactivity of this purified hGM-CSF was measured at an EC₅₀ of 16.4 ± 2 pM by a CCK8 assay in TF-1 human erythroleukemia cells.     

An EM approach to mineral analysis using natural gamma rays

We describe here a method for the analysis of materials on a conveyor belt using the natural gamma spectra collected with a BGO (Bismuth Germanate) gamma ray detector. This detector collects gamma ray emissions from the Potassium (K), Uranium (U), and Thorium (Th) atoms in the materials. Based on these data, and using a Poisson model for the data generation, a statistical model is proposed and an approximate maximum likelihood (ML) technique based on the expectation-maximization (EM) algorithm is then used to estimate the amount of each of the three elements in the material. The statistical model is further refined to incorporate parameters of drift in the detector and an estimation technique for this is developed and tested against real data. The Cramér–Rao lower bounds for the estimators are calculated.     

Short packet communication in underlay cognitive network assisted by an intelligent reflecting surface

We propose short packet communication in an underlay cognitive radio network assisted by an intelligent reflecting surface (IRS) composed of multiple reconfigurable reflectors. This scheme, called the IRS protocol, operates in only one time slot (TS) using the IRS. The IRS adjusts its phases to give zero received cumulative phase at the secondary destination, thereby enhancing the end-to-end signal-to-noise ratio. The transmitting power of the secondary source is optimized to simultaneously satisfy the multi-interference constraints, hardware limitations, and performance improvement. Simulation and analysis results of the average block error rates (BLERs) show that the performance can be enhanced by installing more reconfigurable reflectors, increasing the blocklength, lowering the number of required primary receivers, or sending fewer information bits. Moreover, the proposed IRS protocol always outperforms underlay relaying protocols using two TSs for data transmission, and achieves the best average BLER at identical transmission distances between the secondary source and secondary destination. The theoretical analyses are confirmed by Monte Carlo simulations.     

Wavelet CMAC for MIMO uncertain nonlinear systems using a modified social ski-driver algorithm

This study uses a Mexican hat wavelet membership function for a cerebellar model articulation controller (CMAC) to develop a more efficient adaptive controller for multiple input multiple output (MIMO) uncertain nonlinear systems. The main controller is called the adaptive Mexican hat wavelet CMAC (MWCMAC), and an auxiliary controller is used to remove the residual error. For the MWCMAC, the online learning laws are derived from the gradient descent method. In addition, the learning rate values are very important and have a great impact on the performance of the control system; however, they are difficult to choose accurately. Therefore, a modified social ski driver (SSD) algorithm is proposed to find optimal learning rates for the control parameters. Finally, a magnetic ball levitation system and a nine-link biped robot are used to illustrate the effectiveness of the proposed SSD-based MWCMAC control system. The comparisons with other existing control algorithms have shown the superiority of the proposed control system.     

Intelligent wavelet fuzzy brain emotional controller using dual function-link network for uncertain nonlinear control systems

This study aims to propose a more efficient hybrid algorithm to achieve favorable control performance for uncertain nonlinear systems. The proposed algorithm comprises a dual function-link network-based multilayer wavelet fuzzy brain emotional controller and a sign(.) functional compensator. The proposed algorithm estimates the judgment and emotion of a brain that includes two fuzzy inference systems for the amygdala network and the prefrontal cortex network via using a dual-function-link network and three sub-structures. Three sub-structures are a dual-function-link network, an amygdala network, and a prefrontal cortex network. Particularly, the dual-function-link network is used to adjust the amygdala and orbitofrontal weights separately so that the proposed algorithm can efficiently reduce the tracking error, follow the reference signal well, and achieve good performance. A Lyapunov stability function is used to determine the adaptive laws, which are used to efficiently tune the system parameters online. Simulation and experimental studies for an antilock braking system and a magnetic levitation system are presented to verify the effectiveness and advantage of the proposed algorithm.

     

Workability, mechanical properties, and chemical stability of a recycled tyre rubber-filled cementitious composite

The workability and mechanical properties of mortar containing shredded automobile and truck tyres were evaluated. Two different shapes of rubber particles were used as constituents of mortar: (1) granules about 2 mm in diameter, and (2) shreds having two sizes which were, nominally, 5.5 mm×1.2 mm and 10.8 mm×1.8 mm (length×diameter). As expected, the geometry of the rubber particles influenced the fracture behaviour of rubber-containing mortar. The addition of rubber led to a decrease in flexural strength and plastic shrinkage cracking of mortar. The crack width and crack length due to plastic shrinkage were reduced for mortar containing the 10.8×1.8 mm rubber shreds compared with a mortar without shreds. The rheological properties of the mortar containing rubber shreds were comparable to those of a mortar without rubber and yielded lower plastic viscosity than a mortar containing 25.4 mm×15 m (length×diameter) polypropylene fibres. The alkaline stability of rubber in mortar was also evaluated by immersing rubber shreds in NaOH and Ca(OH)2 solutions for 4 mon and the results showed that there is less than 20% change in stress and strain value. The findings of the research suggest that automobile and truck tyres can be recycled by shredding and incorporating them into mortar and probably concrete for certain infrastructural applications. © 1998 Chapman & Hall     

Hierarchical ATPG for analog circuits and systems

Automatic test-pattern generation (ATPG) algorithms for analog circuits have been under intense investigation for the last several years. As system design aggressively moves to system-on-a-chip (SoC) and core-based integration, hierarchical analog ATPG emerges as an even more difficult challenge. Attempts to develop an effective algorithm have had varying degrees of success. This article reviews some fundamental issues and recent work in hierarchical analog ATPG and presents an algorithm based on controllability and observability computation. This algorithm has been implemented in a prototype tool, and results based on several case studies show the application of the technique