Kinetic properties of a L-cysteine desulfhydrase-deficient mutant in the enzymatic formation of L-cysteine from D,L-ATC

Summary A mutant strain lacking in activity of L-cysteine desulfhydrase, a L-cysteine-decomposing enzyme, was screened after UV-treatment ofPseudomonas sp. CU6. The properties of the two strains, original and mutant, were compared on the basis of parameter values estimated from kinetic simulations of the enzymatic formation of L-cysteine from D,L-ATC. Both strains suffered from product inhibition, though inhibition was less for the mutant strain.     

Work flow control in the flexible flow line

This research involves the development and evaluation of a part flow control model for a type of flexible manufacturing system (FMS) called a dedicated flexible flow line (FFL). In the FFL, all part types flow along the same path between successive machine groups. The specific objective of the part flow control model for the FFL is to minimize makespan for a given set of parts produced in a FFL near-term schedule, given fixed available buffer constraints. The control model developed in this research involved the repeated, real-time execution of a mathematical programming algorithm. The algorithm attempts to release the right mix of parts at the tight time to keep the FFL operating smoothly. The focus of the approach is directed toward managing WIP buffers for each machine group queue. The algorithm specifically incorporates stochastic disturbance factors such as machine failures. Through a limited number of simulation experiments, performance of the control model is shown to be superior to other parts releasing and control methods reported in the literature.     

Analysis of structural polypeptides of purified human cytomegalovirus.

Human cytomegalovirus strain C87 was purified by the following procedures. (i) Extracellular virus was concentrated by centrifugation at 100,000 X g for 90 min and passed through a Bio-Rad Bio-Gel A-15m column. Most of the virus was recovered in the void volume. (ii) After two consecutive isopycnic potassium tartrate gradient centrifugations (20 to 50%), coinciding peaks of plaque titer, protein, and radioactivity were found at a density of from 1.20 to 1.21 g/cm3. To characterize the structural polypeptides of human cytomegalovirus and to establish relative purification criteria, virus was purified from two mixtures: (i) [35S]methionine-labeled extracellular virus mixed with an equal volume of unlabeled normal culture fluid; (ii) unlabeled extracellular virus mixed with an equal volume of [357a1methionine-labeled normal culture fluid. The extent of purification, as judged by the ratio of cellular to viral radioactivity, was 39-fold; i.e. about 2.5% of the protein in the purified virus preparation could be accounted for by host protein contamination. Electrophoresis of purified [35S]methionine-labeled virus on a polyacrylamide gel slab showed that there were at least 33 viral structural polypeptides (VPs), and their molecular weights ranged from 11,000 to 290,000. Autoradiograms obtained from electropherograms of purified [14C]glucosamine labeled virus showed six bands. Four of these were so broad that several VPs corresponded to each of the glycosylated bands. When heavy (two fractions close to 1.21 g/cm3) and light (two fractions close to 1.20 g/cm3) fractions of the PFU peak from the second potassium tartrate gradient were analyzed separately, the number of polypeptides observed was the same, but the relative amounts of some polypeptides differed. The major polypeptide, VP17, was found in greater amounts in the heavy fraction (35%) than in the light fraction (22%). The amount of DNA as a percentage of the weight of protein was 2% for the light fraction and 1% for the heavy fraction.     

Distal NF-kB binding motif functions as an enhancer for nontypeable H. influenzae-induced DEFB4 regulation in epithelial cells

Among the antimicrobial molecules produced by epithelial cells, DEFB4 is inducible in response to proinflammatory signals such as cytokines and bacterial molecules. Nontypeable Haemophilus influenzae (NTHi) is an important human pathogen that exacerbates chronic obstructive pulmonary disease in adult and causes otitis media and sinusitis in children. Previously, we have demonstrated that DEFB4 effectively kills NTHi and is induced by NTHi via TLR2 signaling. The 5′-flanking region of DEFB4 contains several NF-κB binding motifs, but their NTHi-specific activity remains unclear. In this study, we aimed to elucidate molecular mechanism involved in DEFB4 regulation, focusing on the role of the distal NF-κB binding motif of DEFB4 responding to NTHi. Here, we show that the human middle ear epithelial cells up-regulate DEFB4 expression in response to NTHi via NF-κB activation mediated by IκKα/β−IκBα signaling. Deletion of the distal NF-κB binding motif led to a significant reduction in NTHi-induced DEFB4 up-regulation. A heterologous construct containing the distal NF-κB binding motif was found to increase the promoter activity in response to NTHi, indicating a NTHi-responding enhancer activity of the distal NF-κB binding motif. Furthermore, electrophoretic mobility shift assays and chromatin immunoprecipitation assays showed that the p65 domain of NF-κB binds to the distal NF-κB binding motif in response to NTHi. Taken together, our results suggest that NTHi-induced binding of p65 NF-κB to the distal NF-κB binding motif of DEFB4 enhances NTHi-induced DEFB4 regulation in epithelial cells.     

Simulation of sequential batch reactor (SBR) operation for simultaneous removal of nitrogen and phosphorus

Modeling of the operation of sequential batch reactor (SBR) was performed to find out optimum design parameters for simultaneous removal of nitrogen and phosphorus in a small-scale wastewater treatment plant. The models were set up with material balances on SBR operation and Monod kinetics. The model parameters were obtained to best fit the experimental results in a small scale SBR. The models were useful in optimizing hydraulic retention time (HRT) and successfully simulated operations of SBR in a larger scale. Especially the model predicted well the reactions occurring in the filling period as well as the effect of dilution, and evaluated the performance of SBR process under diverse operating conditions.     

Combinatory responses of proinflamamtory cytokines on nitric oxide-mediated function in mouse calvarial osteoblasts

Combinatory responses of proinflamamtory cytokines have been examined on the nitric oxide-mediated function in cultured mouse calvarial osteoblasts. Interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) induced iNOS gene expression and NO production, although these actions were inhibited by L-NG-monomethylarginine (L-NMMA) and decreased alkaline phosphatase (ALPase) activity. Furthermore, NO donors, sodium nitroprusside (SNP) and NONOate dose-dependently elevated ALPase activity. In contrast, transforming-growth factor-β (TGF-β) decreased NO production stimulated by IL-1β, TNF-α and interferon-γ (IFN-γ). iNOS was expressed by mouse calvarial osteoblast cells after stimulation with IL-1β, TNF-α, and IFN-γ. Incubation of mouse calvarial osteoblast cells with the cytokines inhibited growth and ALPase activity. However, TGF-β-treatment abolished these effects of IL-1β, TNF-α and IFN-γ on growth inhibition and stimulation of ALPase in mouse calvarial osteoblast cells. In contrast, IL-1β, TNF-α, and IFN-γ exerted growth-inhibiting effects on mouse calvarial osteoblast cells which were partly NO-dependent. The results suggest that NO may act predominantly as a modulator of cytokine-induced effects on mouse calvarial osteoblast cells and TGF-β is a negative regulator of the NO production stimulated by IL-1β, TNF-α and IFN-γ.