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Yarden Opatowsky Orna Chomsky‐Hecht Joel A. Hirsch 《Acta Crystallographica. Section D, Structural Biology》2004,60(7):1301-1303
Two versions of the functional core of the rabbit voltage‐dependent calcium channel β2a subunit were expressed in Escherichia coli. These proteins were purified to homogeneity and screened for crystallization. Crystallization conditions were refined using the hanging‐drop vapour‐diffusion method and two crystal forms were pursued. Crystal form I is represented by thick rods with tetragonal symmetry, unit‐cell parameters a = b = 75, c = 165 Å and a diffraction limit of 3.4 Å which were obtained using ammonium sulfate as a precipitant. Crystal form II gives rise to plates with orthorhombic symmetry, unit‐cell parameters a = 35, b = 75, c = 165 Å and a diffraction limit of 2.3 Å which were grown using polyethylene glycol 20K as a precipitant. 相似文献
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Huanli Xie Leon Parsaud Jamie A. Lopez Yu He Subbulakshmi Chidambaram Patrick P. Lam David E. James Shuzo Sugita Herbert Y. Gaisano 《Traffic (Copenhagen, Denmark)》2013,14(4):428-439
RalA GTPase has been implicated in the regulated delivery of exocytotic vesicles to the plasma membrane (PM) in mammalian cells. We had reported that RalA regulates biphasic insulin secretion, which we have now determined to be contributed by RalA direct interaction with voltage‐gated calcium (Cav) channels. RalA knockdown (KD) in INS‐1 cells and primary rat β‐cells resulted in a reduction in Ca2+ currents arising specifically from L‐(Cav1.2 and Cav1.3) and R‐type (Cav2.3) Ca2+ channels. Restoration of RalA expression in RalA KD cells rescued these defects in Ca2+ currents. RalA co‐immunoprecipitated with the Cavα2δ‐1 auxiliary subunit known to bind the three Cavs. Moreover, the functional molecular interactions between Cavα2δ‐1 and RalA on the PM shown by total internal reflection fluorescent microscopy/FRET analysis could be induced by glucose stimulation. KD of RalA inhibited trafficking of α2δ‐1 to insulin granules without affecting the localization of the other Cav subunits. Furthermore, we confirmed that RalA and α2δ‐1 functionally interact since RalA KD‐induced inhibition of Cav currents could not be recovered by RalA when α2δ‐1 was simultaneously knocked down. These data provide a mechanism for RalA function in insulin secretion, whereby RalA binds α2δ‐1 on insulin granules to tether these granules to PM Ca2+ channels. This acts as a chaperoning step prior to and in preparation for sequential assembly of exocyst and excitosome complexes that mediate biphasic insulin secretion. 相似文献
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Cheril Tapia‐Rojas Patricia V. Burgos Nibaldo C. Inestrosa 《Journal of neurochemistry》2016,139(6):1175-1191
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Hyunju Kim Yu‐Ran Na So Yeon Kim Eun Gyeong Yang 《Journal of cellular biochemistry》2016,117(3):647-658
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J. L. Tetens S. Qanbari C. Drögemüller E. C. G. Pimentel J. Bennewitz G. Thaller J. Tetens 《Animal genetics》2014,45(4):585-588
The major bovine whey proteins, α‐lactalbumin (α‐LA) and β‐lactoglobulin (β‐LG), exhibit breed‐specific genetic variation. The aim of this study was to identify possible new protein variants and determine the distribution of variants across a variety of 18 taurine and indicine cattle breeds applying a DNA‐based sequencing approach. To this end, the open reading frames of the respective genes (LALBA and LGB) were sequenced in 476 animals. Within the LALBA gene, a previously unknown synonymous and a previously undesignated non‐synonymous nucleotide exchange were identified. Furthermore, two known α‐LA variants (A and B) and four known β‐LG variants (A, B, C and W) were determined. The occurrence of typical indicine variants in some taurine cattle breeds, such as Suisse Eringer, German Hinterwälder and Hungarian Grey Steppe, further supports the hypothesis of ancient Bos indicus introgression into (peri‐)alpine cattle breeds. 相似文献
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《Peptide Science》2017,108(3)
The conformational characteristics of protected homo‐oligomeric Boc‐[β3(R)Val]n‐OMe, n = 1, 2, 3, 4, 6, 9, and 12 have been investigated in organic solvents using nuclear magnetic resonance (NMR), Fourier transform infrared (FTIR) absorption spectroscopy and circular dichroism (CD) methods. The detailed 1H NMR analysis of Boc‐[β3(R)Val]12‐OMe reveals that the peptide aggregates extensively in CDCl3, but is disaggregated in 20%, (v/v) dimethyl sulfoxide (DMSO) in CDCl3 and in CD3OH. Limited assignment of the N‐terminus NH groups, together with solvent dependence of NH chemical shifts and temperature coefficients provides evidence for 14‐helix conformation in the 12‐residue peptide. FTIR analysis in CHCl3 establishes that the onset of folding and aggregation, as evidenced by NH stretching bands at 3375 cm−1 (intramolecular) and 3285 cm−1 (intermolecular), begins at the level of the tetrapeptide. The observed CD bands, 214 nm (negative) and 198 nm (positive), support 14‐helix formation in the 9 and 12 residue sequences. The folding and aggregation tendencies of homo‐oligomeric α‐, β‐, and γ‐ residues is compared in the model peptides Boc‐[ωVal]n‐NHMe, ω = α, β, and γ and n = 1, 2, and 3. Analysis of the FTIR spectra in CHCl3, establish that the tendency to aggregate at the di and tripeptide level follows the order β > α∼γ, while the tendency to fold follows the order γ > β > α. 相似文献
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Zihao Teng Yan Guo Xingqi Liu Jian Zhang Xiaodi Niu Qinlei Yu Xuming Deng Jianfeng Wang 《Journal of cellular and molecular medicine》2019,23(10):6955-6964
Metallo‐β‐lactamases (MBLs) are some of the best known β‐lactamases produced by common Gram‐positive and Gram‐negative pathogens and are crucial factors in the rise of bacterial resistance against β‐lactam antibiotics. Although many types of β‐lactamase inhibitors have been successfully developed and used in clinical settings, no MBL inhibitors have been identified to date. Nitrocefin, checkerboard and time‐kill assays were used to examine the enzyme behaviour in vitro. Molecular docking calculation, molecular dynamics simulation, calculation of the binding free energy and ligand‐residue interaction decomposition were used for mechanistic research. The behaviour of the enzymes in vivo was investigated by a mouse infection experiment. We showed that theaflavin‐3,3´‐digallate (TFDG), a natural compound lacking antibacterial activities, can inhibit the hydrolysis of MBLs. In the checkerboard and time‐kill assays, we observed a synergistic effect of TFDG with β‐lactam antibiotics against methicillin‐resistant Staphylococcus aureus BAA1717. Molecular dynamics simulations were used to identify the mechanism of the inhibition of MBLs by TFDG, and we observed that the hydrolysis activity of the MBLs was restricted by the binding of TFDG to Gln242 and Ser369. Furthermore, the combination of TFDG with β‐lactam antibiotics showed effective protection in a mouse Staphylococcus aureus pneumonia model. These findings suggest that TFDG can effectively inhibit the hydrolysis activity of MBLs and enhance the antibacterial activity of β‐lactam antibiotics against pathogens in vitro and in vivo. 相似文献
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Hannah Scheiblich Anna Schlütter Douglas T. Golenbock Eicke Latz Pilar Martinez‐Martinez Michael T. Heneka 《Journal of neurochemistry》2017,143(5):534-550
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Ann L Cornish Caroline E Sutton Joanne O'Donnell Louise H Cengia Andrew W Roberts Ian P Wicks Kingston H G Mills Ben A Croker 《EMBO reports》2010,11(8):640-646
Reports describing the effect of interferon‐γ (IFNγ) on interleukin‐1β (IL‐1β) production are conflicting. We resolve this controversy by showing that IFNγ potentiates IL‐1β release from human cells, but transiently inhibits the production of IL‐1β from mouse cells. Release from this inhibition is dependent on suppressor of cytokine signalling 1. IL‐1β and Th17 cells are pathogenic in mouse models for autoimmune disease, which use Mycobacterium tuberculosis (MTB), in which IFNγ and IFNβ are anti‐inflammatory. We observed that these cytokines suppress IL‐1β production in response to MTB, resulting in a reduced number of IL‐17‐producing cells. In human cells, IFNγ increased IL‐1β production, and this might explain why IFNγ is detrimental for multiple sclerosis. In mice, IFNγ decreased IL‐1β and subsequently IL‐17, indicating that the adaptive immune response can provide a systemic, but transient, signal to limit inflammation. 相似文献
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Pu Wang Pei‐Pei Guan Jing‐Wen Guo Long‐Long Cao Guo‐Biao Xu Xin Yu Yue Wang Zhan‐You Wang 《Aging cell》2016,15(5):861-871
Cyclooxygenase‐2 (COX‐2) has been recently identified to be involved in the pathogenesis of Alzheimer's disease (AD). Yet, the role of an important COX‐2 metabolic product, prostaglandin (PG) I2, in the pathogenesis of AD remains unknown. Using human‐ and mouse‐derived neuronal cells as well as amyloid precursor protein/presenilin 1 (APP/PS1) transgenic mice as model systems, we elucidated the mechanism of anterior pharynx‐defective (APH)‐1α and pharynx‐defective‐1β induction. In particular, we found that PGI2 production increased during the course of AD development. Then, PGI2 accumulation in neuronal cells activates PKA/CREB and JNK/c‐Jun signaling pathways by phosphorylation, which results in APH‐1α/1β expression. As PGI2 is an important metabolic by‐product of COX‐2, its suppression by NS398 treatment decreases the expression of APH‐1α/1β in neuronal cells and APP/PS1 mice. More importantly, β‐amyloid protein (Aβ) oligomers in the cerebrospinal fluid (CSF) of APP/PS1 mice are critical for stimulating the expression of APH‐1α/1β, which was blocked by NS398 incubation. Finally, the induction of APH‐1α/1β was confirmed in the brains of patients with AD. Thus, these findings not only provide novel insights into the mechanism of PGI2‐induced AD progression but also are instrumental for improving clinical therapies to combat AD. 相似文献