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排序方式: 共有501条查询结果,搜索用时 31 毫秒
1.
Involvement of the Binuclear Copper Site in the Proteolytic Activity of Polyphenol Oxidase 总被引:2,自引:0,他引:2
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) 相似文献
2.
Dissociation between lymphocyte activation for proliferation and for the capacity to adoptively transfer uveoretinitis 总被引:4,自引:0,他引:4
G M Fox T M Redmond B Wiggert T Kuwabara G J Chader I Gery 《Journal of immunology (Baltimore, Md. : 1950)》1987,138(10):3242-3246
We have shown previously that immunization with bovine interphotoreceptor retinoid-binding protein (IRBP) induces in rats severe eye disease, experimental autoimmune uveoretinitis (EAU). This study examined the uveitogenic capacity of IRBP of another species, the monkey, and tested the cross-antigenicity between these two proteins by a battery of immunological assays. Monkey IRBP was found to be approximately 20 times less uveitogenic in Lewis rats than bovine IRBP. High levels of cross-reactivity between bovine and monkey IRBP were demonstrated by antibodies as measured by the enzyme-linked immunosorbent assay, and by the radiometric ear test of delayed-type hypersensitivity, by using rats immunized with either one of the IRBP. On the other hand, lymphocytes from these rats failed to detect the cross-reactivity between the two IRBP by the proliferation response in culture. Yet, such lymphocytes did recognize the nonimmunizing IRBP when activated in culture for acquiring the capacity to adoptively transfer EAU into naive recipients. The data are discussed with regard to the limited usefulness of the lymphocyte proliferation assay for detection of immunopathogenic processes and the role of cross-reacting antigens in initiation of autoimmune responses. 相似文献
3.
Coprophagy was observed in germfree (GF) ICR mice of both sexes, and the results were compared with those of conventional mice. Frequency of coprophagy per animal per day in GF mice was 5.1 in males and 5.8 in females. In conventional (CV) mice, the frequencies were 6.2 in males and 5.3 in females (data from Zoological Science 2:249-255, 1985), with no significant differences compared with GF mice. Coprophagy in CV mice was frequently observed during 6-8 hr after lighting, whereas such close time relationships tended to weaken in GF animals. In a comparison of levels of constituents per unit weight between feces and diet, fecal crude protein and crude fat exhibited lower values than those in the diet. Levels of fecal crude ash and crude fiber were higher than those in the diet, and nitrogen-free extract was almost equal to that in the diet. No essential difference in these tendencies was found compared with CV mice. Levels of fecal vitamin B1, B2, B12 and folic acid were lower than those in the diet. In CV mice, except for vitamin B1, these vitamins exhibited either almost equal or much higher levels compared with those in the diet (data from Experimental Animals 35: 381-386, 1986). From the fact that coprophagy was observed in GF mice, it is suggested that the behavior is inherent in the mouse. 相似文献
4.
Expression and processing of cyanobacterial Mn-stabilizing protein in Escherichia coli 总被引:1,自引:0,他引:1
The woxA gene of cyanobacterium Anacystis nidulans R2, which encodes the precursor of the Mn-stabilizing protein involved in photosynthetic water oxidation, was found to be expressed in Escherichia coli. The 30-kDa expression product was indistinguishable from the authentic mature protein on SDS/PAGE. Upon fractionation of E. coli cells, the expression product was co-precipitated with the membrane fraction, which is consistent with the water-insoluble nature of the authentic mature protein. Analysis of the amino-terminal amino acid sequence of the product revealed that it is identical to the sequence from the 28th residue of the precursor, indicating that the precursor is processed in E. coli. The expression product was digested by trypsinization of E. coli spheroplasts, but not by that of intact cells. This observation suggests that the product is secreted from the cytoplasmic membrane, but not from the outer membrane. The occurrence of both processing and secretion suggests that a signal peptidase of E. coli can recognize the structure for translocation across the thylakoid membrane. Comparison of the signal sequence and the presequence of sweet potato sporamin A suggests that the processing enzymes of the thylakoid membrane and endoplasmic reticulum possess a common substrate specificity. 相似文献
5.
Komeda Yoshibumi; Yamashita Hirofumi; Sato Naoki; Tsukaya Hirokazu; Naito Satoshi 《Plant & cell physiology》1991,32(6):737-743
The 5'-upstream region (2.4 kb) of the gene for phytochromeI from Pisum sativum (phyl) was fused to the uidA gene fromEscherichia coli that encodes ß-glucuronidase (GUS).The resulting PHY-GUS fusion was introduced into Petunia hybridaand was used as a reporter of the expression of the phyI genewhich was recognized by GUS activity. The PHY-GUS fusion wasexpressed at a relatively high level when transgenic plantswere grown in the dark, while leaves and stems of light-grownplants showed background activity. Flowers of light-grown plantswere shown to have significant levels of GUS activity but rootsdid not have such activity. When light-grown transgenic plantswere transferred to the dark, they expressed the activity atlevels that corresponded to those of dark-grown plants. Lighttreatment prior to growth in darkness revealed red/far-red reversibilityof recovery of the activity. Thus, the 2.4-kb fragment fromthe 5' region of the phyI gene carries the information necessaryfor the light-repressible autoregulation. (Received March 30, 1991; Accepted May 20, 1991) 相似文献
6.
Tomizawa Ken-ichi; Ito Naoko; Komeda Yoshibumi; Uyeda Taro Q. P.; Takio Koji; Furuya Masaki 《Plant & cell physiology》1991,32(1):95-102
The pea phytochrome I (PI) cDNA clone, pPP1001, was expressedin E. coli. The plasmid pPP1001 contains pea PI cDNA which coversthe entire coding region with the Shine-Dalgarno consensus sequencejoined upstream of the cDNA in an expression vector pNUT6. ThepPP1001 transformants formed typical inclusion bodies when culturedat 32?C. However, when cultured at 37?C or in the presence ofisopropyl-ß-D-thiogalactopyranoside (IPTG) at 32?C,the bacteria lysed before inclusion body formation. Immuno-stainingwith anti-PI monoclonal antibody, mAP5, of transformants fixedby cold methanol showed that stainable materials were distributedin whole cytoplasmic region. When the inclusion bodies wereobserved clearly, the regions corresponding to the inclusionbodies became difficult to stain. Western blot analysis, however,showed that a ca. 100 kDa PI polypeptide was detected in thefraction from inclusion bodies and a ca. 90 kDa PI polypeptidefrom the soluble fraction. The amino acid sequence analysisof purified 100 kDa PI sample indicated that its amino terminusis blocked. However, minor signals in one experiment yieldeda sequence corresponding to the expected amino terminus of peaPI except for the initiation methionine. One of the anti-peaPI monoclonal antibodies, mAP9, that recognizes the near N-terminusof pea phytochrome was reactive to the 100 kDa polypeptide. (Received June 22, 1990; Accepted November 18, 1990) 相似文献
7.
In order to investigate coprophagy from the viewpoint of nutrition, fecal constituents were analyzed in freeze-dried samples. Feces were collected from 7:00 to 11:00 and from 19:00 to 23:00. Inorganic elements and crude fibers per unit weight were 3-4 times more concentrated in feces than in basal diet, whereas, crude proteins, crude fats and nitrogen-free extract showed various degrees of reduction. There were no differences in these tendencies with sampling time. As for some B vitamins, feces collected from 7:00 to 11:00 contained 22-92% more vitamins than feces collected from 19:00 to 23:00. In comparison with the dietary concentration, vitamin B12 was increased by 124-197 times (520-730 micrograms/100 g) in feces collected between 7:00 and 11:00. Folic acid in feces collected between 7:00 and 11:00 was 10 times greater than that in the diet. On the basis of the findings on vitamins, the effect of a vitamin B12 fortified diet (1,350 micrograms/100 g) on coprophagy was examined. Mean frequency of coprophagy per animal per day was 9.6 when animals were fed on the basal diet, whereas the frequency was immediately and significantly (p less than 0.05 approximately p less than 0.01) reduced to 4.7 after the diet had been replaced by the fortified one. However, coprophagy was not completely inhibited by vitamin B12 fortification. This indicates that some nutrient(s) in feces other than vitamin B12 might be of use to the host, and that otherwise, coprophagy might be a basically habitual form of behavior. Furthermore, under the fortified diet, the frequency of coprophagy increased gradually.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
8.
Samples of human milk obtained from lactating women in the early postpartum period were assayed for oxytocin concentrations by specific RIA, following extraction procedures with Florisil. Mean oxytocin concentrations in human milk at postpartum day 1 to 5 were 4.5 +/- 1.1, 4.7 +/- 1.1, 4.0 +/- 1.3, 3.2 +/- 0.4, 3.3 +/- 0.6 microunits/ml (+/- SE), respectively. Oxytocin levels in milk were significantly increased by nursing (3.1 +/- 0.6, 5.3 +/- 1.0 microunits/ml, respectively). 3H-oxytocin in human milk was stable even after incubation at 37 degrees C for 2 hours. The dilution curve for milk was parallel to the curve for the standard oxytocin. The chromatographic fraction of immunoreactive oxytocin was identical to that of 3H-oxytocin. 3H-oxytocin was administered to lactating rats. Radioactivity in the neonatal gastric contents and plasma were 12.8% and 4.4% of the counts in the maternal plasma. It was made clear that oxytocin is stable in milk and that oxytocin in maternal blood can be transferred to mik and then to neonates. 相似文献
9.
OH-induced free radicals in purine nucleosides and their homopolymers: e.s.r. and spin-trapping with 2-methyl-2-nitrosopropane 总被引:1,自引:0,他引:1
M Kuwabara O Inanami F Sato 《International journal of radiation biology and related studies in physics, chemistry, and medicine》1986,49(5):829-844
Free radicals produced by X-irradiation of N2O-saturated aqueous solutions of purine nucleosides (2'-deoxyadenosine, adenosine, 2'-deoxyguanosine, 3'-deoxyadenosine, guanosine and inosine) and the corresponding homopolymers (poly A and poly I) have been investigated by the technique of spin-trapping and e.s.r. spectroscopy. 2-Methyl-2-nitrosopropane was used as a spin-trap. For 2'-deoxyadenosine and 2'-deoxyguanosine, the resulting spin-adducts were separated by Bio-Gel P-2 column chromatography and analysed by e.s.r. spectroscopy. For homopolymers, e.s.r. spectra were recorded at 50 degrees C after enzymatic digestion to obtain signals with narrower line width. The e.s.r. signal consisting of only a primary triplet without further splittings, which is consistent with assignment to the trapping of an H-abstraction radical at the C4' position of the sugar moiety, was observed in all cases. For 2'-deoxyguanosine an e.s.r. signal consisting of a secondary triplet was observed. Examinations using other spin-trapping reagents such as PBN, 4-PyOBN and DMPO provided no positive evidence supporting the proposal that this was due to an alpha-nitrogen. The e.s.r. signal consisting of a secondary doublet which further splits into a doublet was observed for 2'-deoxyadenosine, adenosine, 3'-deoxyguanosine, 2'-deoxyguanosine, and inosine, and tentatively associated with a radical centered in the sugar moiety. 相似文献
10.
M Kuwabara G Yoshii T Itoh 《International journal of radiation biology and related studies in physics, chemistry, and medicine》1983,44(2):219-224
Free radicals produced by the reactions of hydrated electrons with pyrimidine nucleosides halogenated at the sugar moiety (2'-chloro-2'-deoxyuridine and 2'-chlorothymidine) were studied by e.s.r. and spin-trapping. 2-Methyl-2-nitrosopropane was used as the spin-trap. The usual spin-trapping technique was extended to frozen and deoxygenated systems to avoid contamination of the trapped radicals with side-products by spin-trapping 2-methyl-2-nitrosopropane itself. When this method was applied to 2'-chloro-2'-deoxyuridine, a free radical at the C-2' position of the sugar moiety was spin-trapped together with a free radical at the C-5 position of the base moiety. This indicates that hydrated electrons both add to the base moiety and eliminate halogen anions from the halogenated sugar moiety. In the case of 2'-chlorothymidine, however, only a free radical attributed to H-addition at the C-6 position of the thymine base was observed. No radicals produced by the reaction of hydrated electrons with the halogenated sugar could be spin-trapped. 相似文献