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1.
Salil K. Niyogi Thomas S. Soper Robert S. Foote Frank W. Larimer Richard J. Mural Sankar Mitra Eva H. Lee Richard Machanoff Fred C. Hartman 《Journal of biosciences》1987,11(1-4):203-214
Both Lys-166 and His-291 of ribulosebisphosphate carboxylase/oxygenase fromRhodospirillum rubrum have been implicated as the active-site residue that initiates catalysis. To decide between these two candidates, we resorted
to site-directed mutagenesis to replace Lys-166 and His-291 with several amino acids. All 7 of the position-166 mutants tested
are severely deficient in carboxylase activity, whereas the alanine and serine mutants at position 291 are ∼40% and ∼18% as
active as the native carboxylase, essentially ruling out His-291 in theRhodospirillum rubrum carboxylase (and by inference His-298 in the spinach enzyme) as a catalytically essential residue. The ability of some of
the mutant proteins to undergo carbamate formation or to bind either ribulosebisphosphate or a transition-state analogue remains
largely unimpaired. This implies that Lys-166 is not required for substrate binding; rather, the results corroborate the earlier
postulate that Lys-166 functions as an acid-base group in catalysis or in stabilizing a transition state in the reaction pathway. 相似文献
2.
When monocytes isolated from human blood adhere to glass substratum, actin- and vinculin-containing punctate plaques rapidly appear at the ventral surface of the cells. We show here that highly purified human leukocyte interferon (IFN) can inhibit formation of these adhesion plaques in a dose-dependent manner. Complete inhibition was obtained when 300 IU/ml IFN were added into the cell-seeding medium. Plaques already formed in the absence of IFN were only partially affected by subsequent addition of IFN into the culture medium. Prevention by IFN of the formation of the adhesion plaques was associated with loosened attachment of the cells to the substratum. Effect of IFN on cellular morphology was complex. At higher doses, IFN added to the cultures within 24 h of seeding almost completely inhibited the differentiation of monocytes to macrophages and most of the cells remained rounded. At lower doses, however, an enhancement of the bipolar spreading was seen and the end result was a culture with predominantly elongated fibroblastoid cells. The latter cells, unlike the fibroblastoid cells in untreated monocyte-macrophage cultures, were completely devoid of the actin plaques, while the reorganization of vimentin-type intermediate filaments took place in a normal manner. These results further support the view that the actin- and vinculin-containing plaques have a role in mediating firm adherence of human monocytes to growth substratum. 相似文献
3.
The effect of Type I interferon on bone resorption was studied by measuring its effect on parathormone-stimulated calcium release from neonatal murine calvaria in vitro. A pure human recombinant leukocyte interferon hybrid of the A and D subtypes was used, which has high antiviral activity on mouse cells. Calcium release was inhibited in a dose dependent fashion with 50% inhibition at about 10(-10) M or 600 U/ml, and the inhibition was reversible. The presence of interferon was required before or during the activation phase of the resorptive response, when the formation of osteoclasts from precursor cells would occur. When added to actively resorbing bone it had no effect. The data suggest that Type I interferon can inhibit the parathormone-regulated development of active osteoclasts, possibly by inhibiting osteoclast precursor differentiation. 相似文献
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5.
Danielly Beraldo dos Santos Silva Luana Mireli Carbonera Rodrigues Adriana Araújo de Almeida Kelly Mari Pires de Oliveira Alexéia Barufatti Grisolia/ 《Memórias do Instituto Oswaldo Cruz》2016,111(3):192-199
The azoles are the class of medications most commonly used to fight infections caused
by Candida sp. Typically, resistance can be attributed to mutations
in ERG11 gene (CYP51) which encodes the cytochrome P450
14α-demethylase, the primary target for the activity of azoles. The objective of this
study was to identify mutations in the coding region of theERG11
gene in clinical isolates of Candidaspecies known to be resistant to
azoles. We identified three new synonymous mutations in the ERG11
gene in the isolates of Candida glabrata (C108G, C423T and A1581G)
and two new nonsynonymous mutations in the isolates of Candida
krusei - A497C (Y166S) and G1570A (G524R). The functional consequence of
these nonsynonymous mutations was predicted using evolutionary conservation scores.
The G524R mutation did not have effect on 14α-demethylase functionality, while the
Y166S mutation was found to affect the enzyme. This observation suggests a possible
link between the mutation and dose-dependent sensitivity to voriconazole in the
clinical isolate of C. krusei. Although the presence of the Y166S in
phenotype of reduced azole sensitivity observed in isolate C.
kruseidemands investigation, it might contribute to the search of new
therapeutic agents against resistant Candida isolates. 相似文献
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7.
Φ-Value analysis was used to characterise the structure of the transition state (TS) for folding of POB L146A Y166W, a peripheral subunit-binding domain that folds in microseconds. Helix 2 was structured in the TS with consolidating interactions from the structured loop that connects the two α-helices. This distribution of Φ-values was very similar to that determined for E3BD F166W, a homologue with high sequence and structural similarity. The extrapolated folding rate constants in water at 298 K were 210,000 s− 1 for POB and 27,500 s− 1 for E3BD. A contribution to the faster folding of POB came from its having significantly greater helical propensity in helix 2, the folding nucleus. The folding rate also appeared to be influenced by differences in the sequence and structural properties of the loop connecting the two helices. Unimodal downhill folding has been proposed as a conserved, biologically important property of peripheral subunit-binding domains. POB folds five times faster and E3BD folds slower than a proposed limit of 40,000 s− 1 for barrier-limited folding. However, experimental evidence strongly suggests that both POB L146A Y166W and E3BD F166W fold in a barrier-limited process through a very similar TS ensemble. 相似文献
8.
Audrey Vincent A cha Ouelkdite-Oumouchal Mouloud Souidi Julie Leclerc Bernadette Neve Isabelle Van Seuningen 《World journal of stem cells》2019,11(11):920-936
The recent discovery of cancer cell plasticity, i.e. their ability to reprogram into cancer stem cells (CSCs) either naturally or under chemotherapy and/or radiotherapy, has changed, once again, the way we consider cancer treatment. If cancer stemness is a reversible epigenetic state rather than a genetic identity, opportunities will arise for therapeutic strategies that remodel epigenetic landscapes of CSCs. However, the systematic use of DNA methyltransferase and histone deacetylase inhibitors, alone or in combination, in advanced solid tumors including colorectal cancers, regardless of their molecular subtypes, does not seem to be the best strategy. In this review, we first summarize the knowledge researchers have gathered on the epigenetic signatures of CSCs with the difficulty of isolating rare populations of cells. We raise questions about the relevant use of currently available epigenetic inhibitors (epidrugs) while the expression of numerous cancer stem cell markers are often repressed by epigenetic mechanisms. These markers include the three cluster of differentiation CD133, CD44 and CD166 that have been extensively used for the isolation of colon CSCs.and . Finally, we describe current treatment strategies using epidrugs, and we hypothesize that, using correlation tools comparing associations of relevant CSC markers with chromatin modifier expression, we could identify better candidates for epienzyme targeting. 相似文献
9.
目的:观察跨膜蛋白166(Transmembrane protein 166,TMEM166)基因在小鼠局部脑缺血再灌注损伤(MCAO)后的表达改变及其对脑细胞凋亡的影响。方法:C57/BL6J雄性小鼠50只,体重22-28 g,采用线栓法制作MCAO模型,缺血后动物随机分为再灌注6、12、24、48 h组。采用免疫组化染色的方法观察缺血侧大脑TMEM166、Caspase 3的阳性细胞数目,TUNEL标记法检测细胞凋亡情况,Western blot检测不同再灌注时间点TMEM166、Caspase 3蛋白水平的表达。结果:缺血侧TMEM166的表达随着再灌注时间的延长而增加,24 h达到高峰,48 h后逐渐下降;再灌注6 h后Caspase 3观察到有表达,同样随着再灌注时间而升高,24 h达到高峰。缺血侧细胞凋亡数量变化趋势与TMEM166基本一致,对侧大脑半球上述指标无明显变化。结论:小鼠局部脑缺血再灌注损伤时伴有TMEM166的表达升高,可能通过激活Caspase 3引起脑细胞凋亡。 相似文献
10.
以盐生植物盐穗木为实验材料,从前期构建的高盐胁迫下盐穗木根的小RNA文库中候选了差异表达的miR166a,利用生物信息学从盐穗木转录组数据中预测其靶基因;采用5′RLM-RACE技术鉴定盐穗木miR166a对预测靶基因ATHB8-like的靶向性;通过PCR和RACE技术克隆盐穗木miR166a前体和预测靶基因ATHB8-like全长基因序列,并进行相应的生物信息学分析。结果显示:盐穗木miR166a预测的靶基因为ATHB8-like;通过实验鉴定确实存在靶向切割,具体的切割位点位于miR166a成熟体的14~15碱基之间;miR166a成熟体序列在不同植物中高度保守;克隆获得的盐穗木miR166a前体可折叠成完整的颈环结构,符合miRNA的前体特征,候选植物miR166a前体在进化上没有表现出保守性;预测的靶基因ATHB8-like cDNA全长为2 786bp,开放阅读框为2 526bp,编码841个氨基酸,ATHB8-like具有一个HD-ZIPⅢ结构域,在进化上具有保守性。该研究结果为进一步开展盐穗木miR166a和ATHB8-like的生物学功能奠定了基础。 相似文献