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61.
In pancreatic acini, calcium-mobilizing agents increase intracellular calcium and stimulate the production of diacylglycerol, and then activate protein kinase C (PKC). However, there are few studies which have examined the activation of PKC in intact acini. To examine the activation of PKC in intact acini by calcium-mobilizing agents, we measured the binding of [3H]phorbol-12,13-dibutyrate (PDBu) to intact acini. Acini were incubated with 10 nM [3H]PDBu at 25 degrees C with or without agents. The binding reactions were terminated by filtration. The filters were counted by a scintillation counter after washing. Acini possessed a single class of binding sites to PDBu, with Kd = 70 nM. CCK-8 and carbachol upregulated the binding affinity of PKC to PDBu in the acini. The ability of calcium-mobilizing agents to increase binding of [3H]PDBu to the acini had a close correlation to their ability to stimulate the amylase secretion from the acini, and higher concentrations of CCK-8 for amylase secretion suppressed binding of [3H]PDBu to the acini. 8Br-cAMP, 8Br-cGMP, and calcium ionophore did not inhibit the maximal activation of PKC induced by CCK-8. The calmodulin inhibitor W7 did not reverse the inhibitory effect of higher concentrations of CCK-8 on PKC activation. These results indicate that calcium-mobilizing agents upregulate the binding affinity of PKC to PDBu in intact acini, and that higher concentrations of CCK-8 for amylase secretion may activate the intracellular mechanism that inhibits PKC activity in acini. This inhibitory mechanism was mediated by some other mechanism other than cAMP-, cGMP-, calcium- and calmodulin-dependent mechanisms. 相似文献
62.
F Yanagi T Akasaka S Kozuma K Ichinose K Higashi 《Canadian Metallurgical Quarterly》1998,47(11):1369-1372
Use of a transfusion pump (BP 101, Terumo, Tokyo) makes it feasible to obtain a stable and almost constant ejection volume at a flow rate of 99 ml.min-1, with no untoward effects of the intravenously placed needle and the microfilter located in the circuit. An air sensor ensures that the pump will cease operation immediately and automatically if an air bubble occurrs in the circuit. When a blood bag is conventionally connected to a connecting tube, at the maximum flow rate, one must set up a new blood bag within a few minutes, and in emergency situations with a shortage of hands, this would be unduly troublesome. When 5-7 blood bags (400 ml) is connected to 5-7 parallel connecting tubes, the pump will continuously eject blood approximately to 2000 ml at 99 ml.min-1, without replacing the blood bags. 相似文献
63.
A 4-year-old girl developed numerous tense blisters on the body. The blisters healed without scarring. Histopathological and immunofluorescence studies showed findings consistent with linear immunoglobulin A (IgA) bullous dermatosis of childhood. Immunoelectron microscopy revealed deposition of IgA in the lamina lucida of the basement membrane zone of the dermal-epidermal junction. Circulating IgA autoantibody was positive at the titre of 1 : 128 and recognized the antigens located on epidermal sites of 1 mol/l NaCl-split skin. Immunofluorescence staining of cultured normal human fibroblasts and cultured DJM-1 cells (derived from human squamous cell carcinoma of skin) with the patient's sera demonstrated that both of these cells synthesize the antigens in vitro, although fibroblasts produce the antigens more abundantly. When DJM-1 cells were injected intracutaneously into nude mice, the antigens recognized by the sera were present mainly around the tumour cell islands in a linear pattern, while the dermal-epidermal junction of mouse skin was negative, suggesting that epidermal cells may contribute directly to synthesis and deposition of the antigens at the basement membrane. By immunoprecipitation using cultured normal human fibroblasts, the patient's sera could precipitate at least two specific molecules at 100-kDa and 145-kDa molecular weight. 相似文献
64.
65.
S Higashi T Ohsumi K Ozumi K Kuroki Y Inokuchi M Terashita 《Canadian Metallurgical Quarterly》1998,17(3):186-194
Elimination of the data processing bottleneck in high-throughput sequencing will require both improved accuracy of data processing software and reliable measures of that accuracy. We have developed and implemented in our base-calling program phred the ability to estimate a probability of error for each base-call, as a function of certain parameters computed from the trace data. These error probabilities are shown here to be valid (correspond to actual error rates) and to have high power to discriminate correct base-calls from incorrect ones, for read data collected under several different chemistries and electrophoretic conditions. They play a critical role in our assembly program phrap and our finishing program consed. 相似文献
66.
Hakoshima Toshio; Toda Shoji; Sugio Shigetoshi; Tomita Ken-ichi; Nishikawa Satoshi; Morioka Hiroshi; Fuchimura Kayoko; Kimura Tsuyoshi; Uesugi Sei-ichi; Ohtsuka Eiko; Ikehara Morio 《Protein engineering, design & selection : PEDS》1988,2(1):55-61
Recognition by ribonuclease T1 of guanine bases via multidentatehydrogen bonding and stacking interactions appears to be mediatedmainly by a short peptide segment formed by one stretch of aheptapeptide, Tyr42-Asn43-Asn44-Tyr45-Gly46-Gly47-Phe48. Thesegment displays a unique folding of the polypeptide chainconsistingof a reverse turn, Asn44-Tyr45-Glu46-Gly47, stabilized by ahydrogen-bond network involving the side chain of Asn44, themain-chain atoms of Asn44, Gly47 and Phe48 and one water molecule.The segment is connected to the C terminus of a ß-strandand expands into a loop region between Asn43 and Ser54. Lowvalues for the crystallographic thermal parameters of the segmentindicate that the structure has a rigidity comparable to thatof a ß-pleated sheet. Replacement of Asn44 with alanineleads to a far lower enzymatic activity and demonstrates thatthe side chain of Asn44 plays a key role in polypeptide foldingin addition to a role in maintaining the segment structure.Substitution of Asn43 by alanine to remove a weak hydrogen bondto the guanine base destabilized the transition state of thecomplex by 6.3 kJ/mol at 37°C. In contrast, mutation ofGlu46 to alanine to remove a strong hydrogen bond to the guaninebase caused a destabilization of the complex by 14.0 kJ/mol.A double-mutant enzyme with substitutions of Asn43 by a histidineand Asn44 by an aspartic acid, to reproduce the natural substitutionsfound in ribonuclease Ms, showed an activity and base specificitysimilar to that of the wild-type ribonuclease Ms. The segmenttherefore appears to be well conserved in several fungal ribonucleases. 相似文献
67.
68.
Kinetic study of thermostable L-threonine dehydrogenase from an archaeon Pyrococcus horikoshii 总被引:1,自引:0,他引:1
In the genome data base of the hyperthermophilic archaeon Pyrococcus horikoshii, an open reading frame with sequence homology to a gene encoding alcohol dehydrogenase was found. It was demonstrated that the encoded enzyme was a thermostable L-threonine dehydrogenase which can oxidize the hydroxy alkyl residue of L-threonine associated with the reduction of NAD+ or NADP+. This enzyme is a member of the zinc-containing L-threonine dehydrogenase family. One enzyme molecule contained one zinc atom, and this metal was considered to contribute to the hyperthermostablility of the enzyme. The reaction of the enzyme proceeded via a sequential mechanism. The Michaelis constants (Km) for L-threonine and NAD+ were 0.013 and 0.010 mM, respectively, and the maximum reaction rate (Vmax) was 1.75 mmol NADH formed/min/mg-protein at 65 degrees C. The Km values for both L-threonine and NADP+ were larger than those for L-threonine and NAD+ with a similar Vmax value. These results indicate that the enzyme has lower affinity to NADP+ than to NAD+, and the binding affinity for L-threonine depends on the coenzymes. 相似文献
69.
In this paper, we report the changes in physical properties of polycarbonate caused by absorbed water and supply basic data for the application of this effect. Absorbed water increases monotonously with increase in relative humidity. Along with the increase in absorbed water, the relaxation time of the β process increases, the glass transition temperature rises, the area of the endothermic peak at the glass transition temperature increases, and the dynamic Young's modulus decreases. We conclude that the absorbed water fills holes but does not function as a crosslinking agent. 相似文献
70.