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991.
Ila A Marathe Stella M Lai Walter J Zahurancik Michael G Poirier Vicki
H Wysocki Venkat Gopalan 《Nucleic acids research》2021,49(16):9444
The ribonucleoprotein (RNP) form of archaeal RNase P comprises one catalytic RNA and five protein cofactors. To catalyze Mg2+-dependent cleavage of the 5′ leader from pre-tRNAs, the catalytic (C) and specificity (S) domains of the RNase P RNA (RPR) cooperate to recognize different parts of the pre-tRNA. While ∼250–500 mM Mg2+ renders the archaeal RPR active without RNase P proteins (RPPs), addition of all RPPs lowers the Mg2+ requirement to ∼10–20 mM and improves the rate and fidelity of cleavage. To understand the Mg2+- and RPP-dependent structural changes that increase activity, we used pre-tRNA cleavage and ensemble FRET assays to characterize inter-domain interactions in Pyrococcus furiosus (Pfu) RPR, either alone or with RPPs ± pre-tRNA. Following splint ligation to doubly label the RPR (Cy3-RPRC domain and Cy5-RPRS domain), we used native mass spectrometry to verify the final product. We found that FRET correlates closely with activity, the Pfu RPR and RNase P holoenzyme (RPR + 5 RPPs) traverse different Mg2+-dependent paths to converge on similar functional states, and binding of the pre-tRNA by the holoenzyme influences Mg2+ cooperativity. Our findings highlight how Mg2+ and proteins in multi-subunit RNPs together favor RNA conformations in a dynamic ensemble for functional gains. 相似文献
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Cao YR Wang Q Jin RX Tang SK Jiang Y He WX Lai HX Xu LH Jiang CL 《Antonie van Leeuwenhoek》2011,99(3):629-634
A Gram-negative, pink-pigmented, non-spore-forming rod shaped, methanol-utilizing bacterium, strain YIM 48816(T), was isolated from forest soil collected from Sichuan province, China. Strain YIM 48816(T) can grow at 4-37 °C, pH 5.0-7.0 and 0% NaCl (w/v). Based on 16S rRNA gene sequence similarity studies, it belonged to the genus Methylobacterium, and formed a phyletic line. The 16S rRNA gene sequence similarities were 96.2% to Methylobacterium mesophilicum DSM 1708(T) and 96.0% to Methylobacterium brachiatum DSM 19569(T), and the phylogenetic similarities to all other Methylobacterium species with validly published names were less than 96.0%. The major menaquinones detected were Q-10 (97.14%) and Q-9 (2.86%). The major fatty acids were C18:1 ω7c (80.84%). The DNA G + C content was 66.2 mol%. It is apparent from the genotypic and phenotypic data that strain YIM 48816(T) belongs to a novel species of the genus Methylobacterium, for which the name Methylobacterium soli sp. nov. is proposed. The type strain is YIM 48816(T) (CCTCC AA 208027(T) = KCTC 22810(T)). 相似文献
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Small heat shock proteins, a class of molecular chaperones, are reported to inhibit amyloid fibril formation in vitro, while the mechanism of inhibition remains unknown. In the present study, we investigated the mechanism by which Mj HSP16.5 inhibits amyloid fibril formation of a small peptide (SSTSAA) from RNase A. A model peptide (dansyl-SSTSAA-W) was designed by introducing a pair of fluorescence resonance energy transfer (FRET) probes into the peptide, allowing for the monitoring of fibril formation by this experimental model. Mj HSP16.5 completely inhibited fibril formation of the model peptide at a molar ratio of 1:120. The dynamic process of fibril formation, revealed by FRET, circular dichroism, and electron microscopy, showed a lag phase of about 2 h followed by a fast growth period. The effect of Mj HSP16.5 on amyloid fibril formation was investigated by adding it into the incubation solution during different growth phases. Adding Mj HSP16.5 to the incubating peptide before or during the lag phase completely inhibited fibril formation. However, introducing Mj HSP16.5 after the lag phase only slowed down the fibril formation process by adhering to the already formed fibrils. These findings provide insight into the inhibitory roles of small heat shock proteins on amyloid fibril formation at the molecular level. 相似文献
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Yang S Luo A Hao X Lai Z Ding T Ma X Mayinuer M Shen W Wang X Lu Y Ma D Wang S 《Biology of reproduction》2011,84(6):1182-1189
Peroxiredoxin 2 (PRDX2) has been known to act as an antioxidant enzyme whose main function is H(2)O(2) reduction in cells. We aimed to study the expression patterns of PRDX2 in mouse ovaries and explore the function of this protein in apoptosis of granulosa cells (GCs). We found that the expression of the PRDX2 protein in atretic follicle GCs was markedly higher than in healthy follicle GCs. In vitro, the transfection of siRNA targeting the Prdx2 gene inhibited the proliferation and induced the apoptosis of primary cultured GCs. Furthermore, suppression of PRDX2 resulted in the augmentation of endogenous H(2)O(2), and the ability to eliminate the exogenous H(2)O(2) was attenuated. The expression of PRDX2 and nuclear factor kappa-light-chain-enhancer of activated B cells (NFKB), whose activity was inhibited by binding to IKB, increased in GCs treated with various concentrations of H(2)O(2) for 30 min. However, no significant change in cytoplasmic IKB expression was observed. At 2 h after treatment with H(2)O(2), nuclear NFKB expression level was reduced, cytoplasmic IKB expression was increased, and PRDX2 expression was unchanged. Silencing of the Prdx2 gene caused early changes in NFKB and IKB expression in the primary cultured GCs compared to that in control cells. Taken together, these data suggest that PRDX2 plays an important role in inhibiting apoptosis in GCs and that PRDX2 actions may be related to the expression of NFKB and IKB. 相似文献
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