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321.
Elucidating the ticking of an in vitro circadian clockwork 总被引:1,自引:0,他引:1
Mori T Williams DR Byrne MO Qin X Egli M Mchaourab HS Stewart PL Johnson CH 《PLoS biology》2007,5(4):e93
A biochemical oscillator can be reconstituted in vitro with three purified proteins, that displays the salient properties of circadian (daily) rhythms, including self-sustained 24-h periodicity that is temperature compensated. We analyze the biochemical basis of this oscillator by quantifying the time-dependent interactions of the three proteins (KaiA, KaiB, and KaiC) by electron microscopy and native gel electrophoresis to elucidate the timing of the formation of complexes among the Kai proteins. The data are used to derive a dynamic model for the in vitro oscillator that accurately reproduces the rhythms of KaiABC complexes and of KaiC phosphorylation, and is consistent with biophysical observations of individual Kai protein interactions. We use fluorescence resonance energy transfer (FRET) to confirm that monomer exchange among KaiC hexamers occurs. The model demonstrates that the function of this monomer exchange may be to maintain synchrony among the KaiC hexamers in the reaction, thereby sustaining a high-amplitude oscillation. Finally, we apply the first perturbation analyses of an in vitro oscillator by using temperature pulses to reset the phase of the KaiABC oscillator, thereby testing the resetting characteristics of this unique circadian oscillator. This study analyzes a circadian clockwork to an unprecedented level of molecular detail. 相似文献
322.
Microbiology - In this study, 37 endophytic actinomycetes were isolated from roots of 14 plant species obtained in Thailand, where biological diversity is known to be high. The isolates were... 相似文献
323.
324.
Tsuyoshi Kenri Yoshito Kawakita Hisashi Kudo U. Matsumoto Shigetarou Mori Yukio Furukawa Yuhei O. Tahara Keigo Shibayama Yuuki Hayashi Munehito Arai Makoto Miyata 《Biochemical and biophysical research communications》2019,508(4):1050-1055
Mycoplasma pneumoniae forms an attachment organelle at one cell pole, binds to the host cell surface, and glides via a unique mechanism. A 170-kDa protein, P1 adhesin, present on the organelle surface plays a critical role in the binding and gliding process. In this study, we obtained a recombinant P1 adhesin comprising 1476 amino acid residues, excluding the C-terminal domain of 109 amino acids that carried the transmembrane segment, that were fused to additional 17 amino acid residues carrying a hexa-histidine (6?×?His) tag using an Escherichia coli expression system. The recombinant protein showed solubility, and chirality in circular dichroism (CD). The results of analytical gel filtration, ultracentrifugation, negative-staining electron microscopy, and small-angle X-ray scattering (SAXS) showed that the recombinant protein exists in a monomeric form with a uniformly folded structure. SAXS analysis suggested the presence of a compact and ellipsoidal structure rather than random or molten globule-like conformation. Structure model based on SAXS results fitted well with the corresponding structure obtained with cryo-electron tomography from a closely related species, M. genitalium. This recombinant protein may be useful for structural and functional studies as well as for the preparation of antibodies for medical applications. 相似文献
325.
326.
The human salivary amylase gene (amy1), consisting of eleven exons, is expressed in the salivary gland and in some amylase-producing tumors. Its uppermost exon and the following intron, along with the 5'-flanking region of this gene, are shown to be superimposed with a gamma-actin pseudogene sequence, a portion of which is transcribed into salivary amylase mRNA and another portion of which serves as a promoter for the amy1 gene. In the further upstream region, the gamma-actin pseudogene sequence is interrupted by a human endogenous retroviral nucleotide sequence. 相似文献
327.
1. A kallikrein-like enzyme was isolated and characterized from the venom of Crotalus ruber ruber (red rattlesnake). 2. The kallikrein-like enzyme was shown to be homogeneous as demonstrated by a single band on acrylamide gel electrophoresis, isoelectric focusing, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunodiffusion and reverse-phase (RP) HPLC. 3. The enzyme has a molecular weight of 31,000 and isoelectric point of 4.6. It consists of 271 total amino acid residues, 24% of which are acidic amino acids. 4. Specific esterolytic activities of the kallikrein-like enzyme on N-tosyl-L-arginine methylester (TAME) and N-benzoyl-L-arginine ethylester (BAEE) are 109.5 and 23.6 mumol/min/mg, respectively. 5. The enzyme differs from trypsin as the soybean trypsin inhibitor does not inhibit the enzyme's action. Diisopropylfluorophosphate (DFP) inhibits the enzyme, suggesting that the serine hydroxyl group is important for enzyme activity. 6. The enzyme is not lethal at 15 micrograms/g in mice and has no hemorrhagic activity, yet the injection of the purified enzyme intradermally, produced capillary permeability-increasing activity as shown by the use of Evans blue dye, and immediate drop in blood pressure. It also contracted the rat uterus. 相似文献
328.
Balb/c 3T3 cells have long been used in cell biological studies. When cells were treated with 2 mM sodium butyrate they became flattened and their nuclei became small relative to the cytoplasm. Electron micrographically, intracellular organellae developed significantly: the number of mitochondria increased and membranous structures, namely the rough-surfaced endoplasmic reticulum and Golgi apparatus, developed markedly. Blood coagulation factor VIII was induced in the presence of 2 mM sodium butyrate in a cell sonicate, but not in culture medium. The amount of this factor was almost 100-fold that in the nontreated cells 4 days after culture initiation, indicating that the cloned Balb/c 3T3 cells were of endothelial origin. 相似文献
329.
Aims: The aim of this study is to assess the antibacterial activity of sodium citrate against Streptococcus pneumoniae and several oral bacteria. Methods and Results: The antibacterial activity was determined by broth microdilution method. The results showed that although Enterocuccus faecium OB7084 and Klebsiella pneumoniae OB7088 had high tolerance to sodium citrate, several oral bacteria including Fusobacterium nucleatum JCM8532T, Streptococcus mutans JCM5705T and Strep. pneumoniae NBRC102642T were susceptible. Furthermore, the bactericidal activity of sodium citrate against Strep. pneumoniae NBRC102642T was not influenced by pH in the range of 5·0–8·0, whereas that of sodium lactate was weakened at neutral or weak alkaline pH. When Strep. pneumoniae NBRC102642T was treated with sodium citrate for 2 h, many burst cells were observed. However, addition of MgCl2 or CaCl2 to an assay medium weakened the antimicrobial activity although ZnCl2 or MnCl2 did not influence. Conclusions: Independent of pH, sodium citrate inhibited the growth of oral bacteria, which suggests that the mechanism is different from that of sodium lactate. Significance and Impact of the Study: The results presented in this study would be available for understanding the antimicrobial property of sodium citrate. 相似文献
330.
Disturbed activation of endoplasmic reticulum stress transducers by familial Alzheimer's disease-linked presenilin-1 mutations 总被引:8,自引:0,他引:8
Katayama T Imaizumi K Honda A Yoneda T Kudo T Takeda M Mori K Rozmahel R Fraser P George-Hyslop PS Tohyama M 《The Journal of biological chemistry》2001,276(46):43446-43454
Recent studies have shown independently that presenilin-1 (PS1) null mutants and familial Alzheimer's disease (FAD)-linked mutants should both down-regulate signaling of the unfolded protein response (UPR). However, it is difficult to accept that both mutants possess the same effects on the UPR. Furthermore, contrary to these observations, neither loss of PS1 and PS2 function nor expression of FAD-linked PS1 mutants were reported to have a discernable impact on the UPR. Therefore, re-examination and detailed analyses are needed to clarify the relationship between PS1 function and UPR signaling. Here, we report that PS1/PS2 null and dominant negative PS1 mutants, which are mutated at aspartate residue 257 or 385, did not affect signaling of the UPR. In contrast, FAD-linked PS1 mutants were confirmed to disturb UPR signaling by inhibiting activation of both Ire1alpha and ATF6, both of which are endoplasmic reticulum (ER) stress transducers in the UPR. Furthermore, PS1 mutants also disturbed activation of PERK (PKR-like ER kinase), which plays a crucial role in inhibiting translation during ER stress. Taken together, these observations suggested that PS1 mutations could affect signaling pathways controlled by each of the respective ER-stress transducers, possibly through a gain-of-function. 相似文献