Analysis of Flow Phenomena in Gastric Contents Induced by Human Gastric Peristalsis Using CFD

This paper uses computational fluid dynamics to simulate and analyze intragastric fluid motions induced by human peristalsis. We created a two-dimensional computational domain of the distal stomach where peristalsis occurs. The motion of the gastric walls induced by an antral contraction wave (ACW) on the wall of the computational domain was well simulated using a function defined in this study. Retropulsive flow caused by ACW was observed near the occluded region, reaching its highest velocity of approximately 12 mm/s in the narrowest region. The viscosity of the model gastric contents applied in this study hardly affected the highest velocity, but greatly affected the velocity profile in the computational domain. The shear rate due to gastric fluid motion was calculated using the numerical output data. The shear rate reached relatively high values of approximately 20 s⁻¹ in the most occluded region. The shear rate profile was almost independent of the fluid viscosity. We also simulated mass transfer of a gastric digestive enzyme (pepsin) in model gastric content when peristalsis occurs on the gastric walls. The visualized simulation results suggest that gastric peristalsis is capable of efficiently mixing pepsin secreted from the gastric walls with an intragastric fluid.     

Involvement of the Binuclear Copper Site in the Proteolytic Activity of Polyphenol Oxidase

Plant polyphenol oxidase (PPO) is apt to degrade during andeven after purification. We developed a method to stabilizePPO by 0.3 M NaCl, 0.1% (w/v) Tween 20, and 50% (w/v) ethyleneglycol at pH 6.5. The protein slowly degraded by itself whenthe stabilizing reagents were removed. Ascorbate and/or H₂O₂accelerated the degradation. The ascorbate-induced degradationwas inhibited by catalase, suggesting that H₂O₂ is generatedthrough reduction of PPO by ascorbate. It is likely that dissolvedoxygen is converted to peroxide through two-electron reductionby the reaction center of PPO, binuclear Cu site, and a Fenton-typereaction occurred on it. This understanding was supported bythe finding that the H₂O₂-induced degradation was inhibitedby metal-chelators as well as by polyphenolic substrate of PPO.Considering the postulated mechanism of the self-degradationof PPO, we re-examined the degradation of the 23-kDa proteinof PSII by PPO [Kuwabara et al. (1997) Plant Cell Physiol. 38:179]. The obtained results suggested that the 23-kDa proteintriggers the active oxygen production by the binuclear Cu site,probably as reductant, and receives the radical species preferentiallyto the polypeptide moiety of PPO. (Received April 15, 1999; Accepted July 21, 1999)     

Crystal structure of human PACRG in complex with MEIG1 reveals roles in axoneme formation and tubulin binding

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Electron paramagnetic resonance study of ferrous cytochrome P-450scc-nitric oxide complexes: effects of cholesterol and its analogues

Electron paramagnetic resonance (EPR) spectra of nitric oxide (NO) complexes of ferrous cytochrome P-450scc were measured at 77 K for the first time without using the rapid-mixing and freeze-quenching technique. Without substrate the EPR spectra were very similar to those of cytochrome P-450cam (from Pseudomonas putida) and cytochrome P-450LM (from rat liver microsomes) with rhombic symmetry; gx = 2.071, gz = 2.001, gy = 1.962, and Az = 2.2 mT for 14NO complexes. Upon addition of substrates [such as cholesterol, 22(S)-hydroxycholesterol, 22(R)-hydroxycholesterol, 25-hydroxycholesterol, and 22-ketocholesterol], the EPR spectra exhibited many variations having rhombic symmetry in the major component and an additional minor component with less rhombic symmetry. Furthermore, addition of 20(S)-hydroxycholesterol caused a striking change in the EPR spectrum. The component with rhombic symmetry disappeared completely, and the component with less rhombic symmetry dominated (gx = 2.027, gz = 2.007, gy = 1.984, and Az = 1.76 mT for 14NO complexes). These observations suggest the existence of the following physiologically important natures: (1) the conformational flexibility of the active site of the enzyme due to the steric interaction between the substrate and the heme-bound ligand molecule and (2) the importance of the hydroxylation of the cholesterol side chain at the 20S position to proceed the side-chain cleavage reaction in cytochrome P-450scc.     

The effects of deuterium oxide (2H2O) on the polymerization of tubulin in vitro

The initial rate and final extent of polymerization of both bovine brain tubulin and sea urchin egg tubulin were enhanced in the presence of ²H₂O. The yields were increased in association with the elevation of the ²H₂O concentration. ²H₂O also reduced the critical concentration for polymerization of brain tubulin. Thermodynamic analysis was attempted using the temperature dependence of the critical concentration for polymerization in the presence of ²H₂O. We obtained linear van 't Hoff plots and calculated thermodynamic parameters which were positive and were increased with the elevation of the ²H₂O concentration. The enhancement of the polymerization of tubulin by ²H₂O could, therefore, be the result of the strenghening of intra-and/or inter-molecular hydrophobic interactions of the tubulin molecules. We believe that the increase in lenghth and number of microtubules of the mitotic spindles in the dividing cells of the eukaryotes with ²H₂O may be caused by the direct involvement of ²H₂O in the polymerization of tubulin.     

Existence of multiple forms of cytochrome P-450scc purified from bovine adrenocortical mitochondria

Three fractions of cytochrome P-450scc (denoted as fractions a, b, and c) were purified by a new procedure from bovine adrenocortical mitochondria. The amino-acid content analyses of these three fractions showed no difference. NH2-terminal amino-acid sequences of cytochrome P-450scc fractions, a and b agreed completely with the sequence deduced by nucleotide sequence of cDNA of cytochrome P-450scc mRNA (Morohashi, K., Fujii-Kuriyama, Y., Okada, Y., Sogawa, K., Hirose, T., Inayama, S. and Omura, T. (1984) Proc. Natl. Acad. Sci. USA 81, 4647-4651), whereas the sequence of fraction c showed a missing of isoleucine at the NH2-terminal. COOH-terminal ámino-acid sequences of fractions a, b and c were -Gln-Ala-COOH, identical with the deduced sequence from the cDNA. Measurements of the enzymatic activities of cholesterol side-chain cleavage reaction revealed no distinct difference among these three fractions. Although each of these fractions appeared as a single protein staining band upon SDS-polyacrylamide gel electrophoresis, these fractions showed heterogeneities upon two-dimensional electrophoresis and chromatofocusing. Fraction a contained the major form of cytochrome P-450scc, and its isoelectric point was estimated to be pH 7.8 by isoelectric focusing under both native and denatured conditions, and this value was confirmed by chromatofocusing. Neither of the carbohydrate-specific stainings (such as periodic acid-Schiff staining and lectin-peroxidase stainings using concanavalin A, wheat-germ agglutinin, and soybean agglutinin) of purified cytochrome P-450scc fractions after the electrophoretic resolution on SDS-polyacrylamide gel could show cytochrome P-450scc fractions as glycoproteins, suggesting that the heterogeneities were not due to the glycosylation state.     

Induction of histidine decarboxylase by dexamethasone in mastocytoma P-815 cells

Dexamethasone at a concentration as low as 10 nM significantly increased both the histamine content and histidine decarboxylase activity of cultured mastocytoma P-815 cells. Both effects were clearly seen using several glucocorticoids, which were as effective as dexamethasone. In contrast to that of histamine, the serotonin level of mastocytoma P-815 cells was decreased by treatment with dexamethasone. The dexamethasone-induced increases in histamine content and histidine decarboxylase activity were completely suppressed by the addition of cycloheximide and actinomycin D. Mastocytoma P-815 cells were found to possess binding sites for [3H]dexamethasone in the cytosol (Kd = 15.7 nM) and the nuclei (Kd = 1.26 nM). These results show that glucocorticoids significantly stimulate de novo synthesis of histidine decarboxylase.     

Electrophoretic Studies on Plant Protoplasts I. pH Dependence of Zeta Potentials of Protoplasts from Various Sources

Electrophoretic mobilities of plant protoplasts from varioussources were measured, as a function of the pH of the medium,by a micro-electrophoresis technique to characterize the protoplastsin terms of curves of zeta potential vs. protoplast surfacepH (pH_s). The shape of the curves of zeta potential vs. pH_scurves differed among preparations of protoplasts isolated fromvarious species and strains. The isoelectric points (pI) ofthe protoplasts measured in this study were between 3.0 and4.0. These differences among the protoplasts suggest that itmay be possible to develop an electrophoretic method for theselection of protoplasts. The shape of the curves of zeta potentialvs. pH_s also indicated that carboxyl groups, as well as phosphategroups, may contribute to the negative charges on the surfaceof protoplasts. (Received October 14, 1988; Accepted February 22, 1989)